Compositions and methods for treating estrogen-related medical disorders

ABSTRACT

Disclosed is a compound of formula (I). or a pharmaceutically acceptable salt thereof. Also disclosed are pharmaceutical compositions including the compound of formula (I) and methods of using the compound of formula (I).

CROSS-REFERENCE TO RELATED APPLICATIONS

This is a continuation of U.S. patent application Ser. No. 14/438,361,filed on Apr. 24, 2015, which is a U.S. national stage entry ofInternational Patent Application No. PCT/US2013/066702, filed on Oct.24, 2013, which claims priority to U.S. Provisional Patent ApplicationNo. 61/809,101, filed on Apr. 5, 2013, and U.S. Provisional PatentApplication No. 61/718,035, filed on Oct. 24, 2012, the entire contentsof all of which are fully incorporated herein by reference.

STATEMENT OF GOVERNMENT INTEREST

This invention was made with government support under contract numberNIH RO1 CA102590 awarded by the National Institutes of Health. Thegovernment has certain rights in the invention.

TECHNICAL FIELD

The present invention relates to compounds, compositions, and methodsfor treating and identifying estrogen-related medical disorders such ascancer, inflammation, osteoporosis, vaginal atrophy, central nervousdiseases, and cardiovascular system diseases.

BACKGROUND

In addition to developmental functions, estrogens have been found toreduce incidence of coronary heart disease (1), maintain bone mineraldensity (2), and in the CNS, promote neuronal survival (3, 4) andhippocampal neurogenesis (5-7). Neuro-imaging studies reveal thatestrogen therapy improves cerebral blood flow and performance inhippocampal-dependent memory tasks in humans (8, 9). Other observationalstudies have found that estrogen helps alleviate age-related cognitivedecline by preserving executive function in the frontal lobe (10).

Raloxifene (EVISTA) is a second generation selective estrogen receptormodulator (SERM) used clinically for the treatment of osteoporosis inpostmenopausal women, which acts as an antiestrogen in breast andendometrium (17-21). Raloxifene, however, increases the lifetime risk ofthromboembolism (23). Raloxifene has been found to enhance levels ofvasodilatory NO through action on endothelial nitric oxide synthase(eNOS) (24-26), and, the increased thromboembolic events have beenattributed to decreased eNOS activity in postmenopausal women (27). NOinhibits thrombus formation through inhibition of platelet recruitment,adhesion and aggregation (28). The next generation SERM, arzoxifene (41,42), a prodrug of desmethylarzoxifene (DMA) was designed to improve thepharmacokinetic (PK) profile relative to raloxifene, from which itdiffers by only one atom (43, 44).

SUMMARY

In one aspect, disclosed is a compound of formula (I)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. X₁ isselected from the group consisting of alkyl, —O—, —N(H)—, —S—, —S(═O)—,and —C(═O)—. A₃ is selected from the group consisting of alkyl, aryl,cycloalkyl, cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selectedfrom the group consisting of hydrogen, halogen, alkyl, cyano,trifluoromethyl, aryl-heteroaryl wherein the aryl is substituted orunsubstituted (e.g., 1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃.R₁, R₂ and R₃ are each independently selected from the group consistingof hydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and—R₄-G₁. R^(x1), R^(y1) and R^(z1), at each occurrence, are independentlyselected from the group consisting of hydrogen and a pharmaceuticallyacceptable cation. R^(a) is alkyl or —OH. R₄, at each occurrence, isindependently selected from the group consisting of a substituentcomprising 1 to 10 carbon atoms, a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen atom, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one C═O group, and a substituent comprising 1 to 10 carbon atomsand optionally containing at least one nitrogen and/or at least one C═Ogroup. G₁, at each occurrence, is independently selected from the groupconsisting of hydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃is —R₄-G₁, and wherein at least one occurrence of G₁ is —ONO₂.

Also provided herein is a compound of formula (I), wherein R₄, at eachoccurrence, is independently selected from the group consisting ofalkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

Also provided herein is a compound of formula (I), wherein R₄, at eachoccurrence, is independently selected from the group consisting ofalkyl, alkylheterocyclealkyl, and heterocyclealkyl.

Also provided herein is a compound of formula (I), wherein A₄ isselected from the group consisting of halogen and —OR₃.

The compound of formula (I) may comprise a compound of formula (I-i), acompound of formula (I-ii), a compound of formula (I-iii), a compound offormula (I-iv), a compound of formula (I-v), or a compound of formula(I-vi)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. X₁ isselected from the group consisting of alkyl, —O—, —N(H)—, —S—, —S(═O)—,and —C(═O)—. A₃ is selected from the group consisting of alkyl, aryl,cycloalkyl, cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selectedfrom the group consisting of hydrogen, halogen, alkyl, cyano,trifluoromethyl, aryl-heteroaryl wherein the aryl is substituted orunsubstituted (e.g., 1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃.R₁, R₂ and R₃ are each independently selected from the group consistingof hydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and—R₄-G₁. R^(x1), R^(y1) and R^(z1), at each occurrence, are independentlyselected from the group consisting of hydrogen and a pharmaceuticallyacceptable cation. R^(a) is alkyl or —OH. R₄, at each occurrence, isindependently selected from the group consisting of a substituentcomprising 1 to 10 carbon atoms, a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen atom, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one C═O group, and a substituent comprising 1 to 10 carbon atomsand optionally containing at least one nitrogen and/or at least one C═Ogroup. G₁, at each occurrence, is independently selected from the groupconsisting of hydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃is —R₄-G₁, and wherein at least one occurrence of G₁ is —ONO₂.

Also provided herein is a compound of formula (I-i), a compound offormula (I-ii), a compound of formula (I-iii), a compound of formula(I-iv), a compound of formula (I-v), or a compound of formula (I-vi),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

Also provided herein is a compound of formula (I-i), a compound offormula (I-ii), a compound of formula (I-iii), a compound of formula(I-iv), a compound of formula (I-v), or a compound of formula (I-vi),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. R₁ and R₂ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R₄, at eachoccurrence, is independently selected from the group consisting of asubstituent comprising 1 to 10 carbon atoms, a substituent comprising 1to 10 carbon atoms and optionally containing at least one nitrogen atom,a substituent comprising 1 to 10 carbon atoms and optionally containingat least one C═O group, and a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen and/or at leastone C═O group. R₅ is selected from the group consisting of hydrogen andC₁-C₃-alkyl-G₁. G₁, at each occurrence, is independently selected fromthe group consisting of hydrogen and —ONO₂, wherein at least one of R₁and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁, and wherein at least oneoccurrence of G₁ is —ONO₂. R^(x1), R^(y1) and R^(z1) are eachindependently selected from the group consisting of hydrogen and apharmaceutically acceptable cation. R^(a) is alkyl or —OH.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia),wherein —R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ia-i), acompound of formula (Ia-ii), a compound of formula (Ia-iii), a compoundof formula (Ia-iv), a compound of formula (Ia-v), or a compound offormula (Ia-vi)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. R₁ and R₂ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R₄, at eachoccurrence, is independently selected from the group consisting of asubstituent comprising 1 to 10 carbon atoms, a substituent comprising 1to 10 carbon atoms and optionally containing at least one nitrogen atom,a substituent comprising 1 to 10 carbon atoms and optionally containingat least one C═O group, and a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen and/or at leastone C═O group. R₅ is selected from the group consisting of hydrogen andC₁-C₃-alkyl-G₁. G₁, at each occurrence, is independently selected fromthe group consisting of hydrogen and —ONO₂, wherein at least one of R₁and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁, and wherein at least oneoccurrence of G₁ is —ONO₂. R^(x1), R^(y1) and R^(z1) are eachindependently selected from the group consisting of hydrogen and apharmaceutically acceptable cation. R^(a) is alkyl.

The compound of formula (I) may comprise a compound of formula (Ia-i), acompound of formula (Ia-ii), a compound of formula (Ia-iii), a compoundof formula (Ia-iv), a compound of formula (Ia-v), or a compound offormula (Ia-vi), wherein R₄, at each occurrence, is independentlyselected from the group consisting of alkyl, cycloalkyl,cycloalkylalkyl, heterocyclealkyl, and alkylheterocyclealkyl, whereinany carbon atom on the alkyl group, together with an alkylene, may forma cycloalkyl, and wherein alkyl may be unsubstituted or substituted with1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ia-i), acompound of formula (Ia-ii), a compound of formula (Ia-iii), a compoundof formula (Ia-iv), a compound of formula (Ia-v), or a compound offormula (Ia-vi), wherein R₄, at each occurrence, is independentlyselected from the group consisting of alkyl, alkylheterocyclealkyl, andheterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia-i), acompound of formula (Ia-ii), a compound of formula (Ia-iii), a compoundof formula (Ia-iv), a compound of formula (Ia-v), or a compound offormula (Ia-vi), wherein —R₄-G₁ is

The compound of formula (I) may comprise formula (Ib)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. R₁ andR₂ are each independently selected from the group consisting ofhydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R⁴-G₁.R₄, at each occurrence, is independently selected from the groupconsisting of a substituent comprising 1 to 10 carbon atoms, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen atom, a substituent comprising 1 to 10 carbon atomsand optionally containing at least one C═O group, and a substituentcomprising 1 to 10 carbon atoms and optionally containing at least onenitrogen and/or at least one C═O group. R₅ is selected from the groupconsisting of hydrogen and C₁-C₃-alkyl-G₁. G₁, at each occurrence, isindependently selected from the group consisting of hydrogen and —ONO₂,wherein at least one of R₁ and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁,and wherein at least one occurrence of G₁ is —ONO₂. R^(x1), R^(y1) andR^(z1) are each independently selected from the group consisting ofhydrogen and a pharmaceutically acceptable cation. R^(a) is alkyl or—OH.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ib-i), acompound of formula (Ib-ii), a compound of formula (Ib-iii), a compoundof formula (Ib-iv), a compound of formula (Ib-v), or a compound offormula (Ib-vi)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. R₁ andR₂ are each independently selected from the group consisting ofhydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R⁴-G₁.R₄, at each occurrence, is independently selected from the groupconsisting of a substituent comprising 1 to 10 carbon atoms, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen atom, a substituent comprising 1 to 10 carbon atomsand optionally containing at least one C═O group, and a substituentcomprising 1 to 10 carbon atoms and optionally containing at least onenitrogen and/or at least one C═O group. R₅ is selected from the groupconsisting of hydrogen and C₁-C₃-alkyl-G₁. G₁, at each occurrence, isindependently selected from the group consisting of hydrogen and —ONO₂,wherein at least one of R₁ and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁,and wherein at least one occurrence of G₁ is —ONO₂. R^(x1), R^(y1) andR^(z1) are each independently selected from the group consisting ofhydrogen and a pharmaceutically acceptable cation. R^(a) is alkyl or—OH.

The compound of formula (I) may comprise a compound of formula (Ib-i), acompound of formula (Ib-ii), a compound of formula (Ib-iii), a compoundof formula (Ib-iv), a compound of formula (Ib-v), or a compound offormula (Ib-vi), wherein R₄, at each occurrence, is independentlyselected from the group consisting of alkyl, cycloalkyl,cycloalkylalkyl, heterocyclealkyl, and alkylheterocyclealkyl, whereinany carbon atom on the alkyl group, together with an alkylene, may forma cycloalkyl, and wherein alkyl may be unsubstituted or substituted with1, 2, or 3 oxo substituents.

The compound of formula (I) may a compound of formula (Ib-i), a compoundof formula (Ib-ii), a compound of formula (Ib-iii), a compound offormula (Ib-iv), a compound of formula (Ib-v), or a compound of formula(Ib-vi), wherein R₄, at each occurrence, is independently selected fromthe group consisting of alkyl, alkylheterocyclealkyl, andheterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ib-i), acompound of formula (Ib-ii), a compound of formula (Ib-iii), a compoundof formula (Ib-iv), a compound of formula (Ib-v), or a compound offormula (Ib-vi), wherein R₄-G₁ is

The compound of formula (I) may comprise formula (Ic)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R^(x1), R^(y1)and R^(z1), at each occurrence, are independently selected from thegroup consisting of hydrogen and a pharmaceutically acceptable cation.R^(a) is alkyl or —OH. R₄, at each occurrence, is independently selectedfrom the group consisting of a substituent comprising 1 to 10 carbonatoms, a substituent comprising 1 to 10 carbon atoms and optionallycontaining at least one nitrogen atom, a substituent comprising 1 to 10carbon atoms and optionally containing at least one C═O group, and asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen and/or at least one C═O group. G₁, at eachoccurrence, is independently selected from the group consisting ofhydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃ is —R₄-G₁, andwherein at least one occurrence of G₁ is —ONO₂.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ic-i), acompound of formula (Ic-ii), a compound of formula (Ic-iii), a compoundof formula (Ic-iv), a compound of formula (Ic-v), or a compound offormula (Ic-vi)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R^(x1), R^(y1)and R^(z1), at each occurrence, are independently selected from thegroup consisting of hydrogen and a pharmaceutically acceptable cation.R^(a) is alkyl or —OH. R₄, at each occurrence, is independently selectedfrom the group consisting of a substituent comprising 1 to 10 carbonatoms, a substituent comprising 1 to 10 carbon atoms and optionallycontaining at least one nitrogen atom, a substituent comprising 1 to 10carbon atoms and optionally containing at least one C═O group, and asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen and/or at least one C═O group. G₁, at eachoccurrence, is independently selected from the group consisting ofhydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃ is —R₄-G₁, andwherein at least one occurrence of G₁ is —ONO₂.

The compound of formula (I) may comprise a compound of formula (Ic-i), acompound of formula (Ic-ii), a compound of formula (Ic-iii), a compoundof formula (Ic-iv), a compound of formula (Ic-v), or a compound offormula (Ic-vi), wherein R₄, at each occurrence, is independentlyselected from the group consisting of alkyl, cycloalkyl,cycloalkylalkyl, heterocyclealkyl, and alkylheterocyclealkyl, whereinany carbon atom on the alkyl group, together with an alkylene, may forma cycloalkyl, and wherein alkyl may be unsubstituted or substituted with1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ic-i), acompound of formula (Ic-ii), a compound of formula (Ic-iii), a compoundof formula (Ic-iv), a compound of formula (Ic-v), or a compound offormula (Ic-vi), wherein R₄, at each occurrence, is independentlyselected from the group consisting of alkyl, alkylheterocyclealkyl, andheterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ic-i), acompound of formula (Ic-ii), a compound of formula (Ic-iii), a compoundof formula (Ic-iv), a compound of formula (Ic-v), or a compound offormula (Ic-vi), wherein R₄-G₁ is

The compound of formula (I) may be selected from the group consistingof:

-   3-(1-(2-(4-((6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-2-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(1-(2-(4-((6-hydroxy-3-(4-hydroxyphenyl)benzo[b]thiophen-2-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-3-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(4-fluorophenyl)-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate; and-   (1-(((2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   or a pharmaceutically acceptable salt thereof.

In another aspect, disclosed is a compound of formula (II),

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1) and —C(═O)R^(a). R^(x1), R^(y1) and R^(z1),at each occurrence, are independently selected from the group consistingof hydrogen and a pharmaceutically acceptable cation. R^(a) is alkyl or—OH.

Also provided are compounds of formula (II), wherein A₁ is fluoro. A₁may be fluoro located in the 4-position of the phenyl ring to which itis attached. Also provided are compounds of formula (II), wherein A₂ is—OH.

The compound of formula (II) may comprise a compound of formula (II-i),a compound of formula (II-ii), a compound of formula (II-iii), acompound of formula (II-iv), a compound of formula (II-v), or a compoundof formula (II-vi)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), and —C(═O)R^(a). R^(x1), R^(y1) andR^(z1), at each occurrence, are independently selected from the groupconsisting of hydrogen and a pharmaceutically acceptable cation. R^(a)is alkyl or —OH.

Also provided herein is a compound of formula (II-i), a compound offormula (II-ii), a compound of formula (II-iii), a compound of formula(II-iv), a compound of formula (II-v), or a compound of formula (II-vi),wherein A₁ is fluoro. A₁ may be fluoro located in the 4-position of thephenyl ring to which it is attached. Also provided herein is a compoundof formula (II-i), a compound of formula (II-ii), a compound of formula(II-iii), a compound of formula (II-iv), a compound of formula (II-v),or a compound of formula (II-vi), wherein A₂ is —OH.

The compound of formula (II) may be selected from the group consistingof:

-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;-   cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;-   1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;-   1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;-   (4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(p-tolyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;    and-   (3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   or a pharmaceutically acceptable salt thereof.

In another aspect, disclosed is a method for treatment of anestrogen-related medical disorder. The method may comprise administeringto a subject in need of such treatment a composition comprising atherapeutically effective amount of at least one compound of formula (I)or a pharmaceutically acceptable salt thereof. The method may compriseadministering to a subject in need of such treatment a compositioncomprising a therapeutically effective amount of at least one compoundof formula (II) or a pharmaceutically acceptable salt thereof.

The estrogen-related medical disorder may be selected from the groupconsisting of: cancer, inflammation, osteoporosis, vaginal atrophy,central nervous system diseases, and cardiovascular system diseases. Thecentral nervous system disease may be selected from the group consistingof Alzheimer's Disease and mild cognitive impairment. The cardiovasculardisease may be thrombosis. The cancer may be selected from the groupconsisting of breast cancer, ovarian cancer, prostate cancer, and lungcancer. The breast cancer may be a tamoxifen resistant breast cancer ora triple negative breast cancer.

The at least one compound of formula (I) may be selected from the groupconsisting of:3-(1-(2-(4-((6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate;4-(6-hydroxy-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-2-yl)phenyl1-((nitrooxy)methyl)cyclopropanecarboxylate;2-(4-fluorophenyl)-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;3-(1-(2-(4-((6-hydroxy-3-(4-hydroxyphenyl)benzo[b]thiophen-2-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate;4-(6-hydroxy-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-3-yl)phenyl1-((nitrooxy)methyl)cyclopropanecarboxylate;3-(4-fluorophenyl)-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; and pharmaceutically acceptable salts thereof.

The at least one compound of formula (II) may be selected from the groupconsisting of:(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;(4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(p-tolyl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;and pharmaceutically acceptable salts thereof.

In another aspect, disclosed is a method of identifying a cancer in asubject. The method may comprise obtaining a test sample from thesubject having cancer, and determining an amount of protein kinase Calpha (PKCα) in the test sample. If the amount of PKCα in the testsample is greater than an amount of PKCα in a test sample from a subjectnot having cancer, then the cancer may be sensitive to at least onecompound of formula (I) or a pharmaceutically acceptable salt thereof,or at least one compound of formula (II) or a pharmaceuticallyacceptable salt thereof.

The at least one compound of formula (I) may be selected from the groupconsisting of:3-(1-(2-(4-((6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate;4-(6-hydroxy-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-2-yl)phenyl1-((nitrooxy)methyl)cyclopropanecarboxylate;2-(4-fluorophenyl)-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;3-(1-(2-(4-((6-hydroxy-3-(4-hydroxyphenyl)benzo[b]thiophen-2-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate;4-(6-hydroxy-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-3-yl)phenyl1-((nitrooxy)methyl)cyclopropanecarboxylate;3-(4-fluorophenyl)-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; 3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate;2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl1-((nitrooxy)methyl)cyclopropanecarboxylate;(1-(((2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methylnitrate; and pharmaceutically acceptable salts thereof.

The at least one compound of formula (II) may be selected from the groupconsisting of:(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;(4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(p-tolyl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;and pharmaceutically acceptable salts thereof.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows SERM-elicited neuroprotection in primary cortical neuronsexposed to OGD is GPR30 dependent. Primary neuronal cultures weresubjected to 2 h OGD with compounds added at the start of OGD andinhibitors added 45 min prior to OGD. Cell survival was measured at 24h. Use of pathway-selective inhibitors indicates that neuroprotection ofDMA and NO-DMA is mediated through PI3K-dependent GPR30 signaling in anER- and NOS-independent manner. Data show mean and S.E.M. normalized toveh. control and E2 (n=6); **=p<0.01, ***=p<0.001 compared to veh.control using one-way ANOVA with Dunnett's post hoc test.

FIG. 2 shows reversal of LTP deficit in aged 3×Tg mice by SERMs is GPR30dependent. LTP was measured after TBS in the CA1 region of hippocampalsections from 16 month male 3×Tg mice or WT controls. Test compounds(SERMs 100 nM; G15 100 nM) were added 30 min prior to TBS. (A, B) DMAand NO-DMA restored deficits in LTP to WT levels and G15 blocked theactions of DMA. (C) Secondary analysis of theta bursts indicate actionboth during induction and stabilization of LTP, through a GPR30dependent mechanism. Data show mean and S.E.M. normalized to baseline(n=4-9); *=p<0.05, **=p<0.01 compared to transgenic veh. control usingone-way ANOVA with Dunnett's post hoc test.

FIG. 3 shows relaxation of isolated aortic rings induced by SERMs andNO-SERM. (A) The EC₅₀ values for relaxation were not significantlydifferent (p>0.05, one-way ANOVA and Newman-Keuls post-hoc test). Themaximal relaxation responses for arzoxifene and FDMA were significantlyless than those for DMA and raloxifene (p<0.05, one-way ANOVA andNewman-Keuls post-hoc test). Each value represents the mean±S.E.M.(n=7-13). (B) Removal of the endothelium or inhibition of NOS withL-NAME reduced the maximal relaxation response to DMA (p<0.01, one-wayANOVA and Newman-Keuls post-hoc test). Each value represents themean±S.E.M. (n=7). (C) The EC₅₀ values for relaxation were significantlyincreased in the presence of L-NAME or after endothelium removal(p<0.05, one-way ANOVA and Newman-Keuls post-hoc test). Each valuerepresents the mean±S.E.M. (n=7).

FIG. 4 shows effects of SERM and NO-SERMon NO levels in plasma and CNSof WT and eNOS (−/−) mice. Levels of NO were assessed by measuringbreakdown products 1 h after i.p. injection of SERMs (2 mg/kg) usingchemiluminescence detection. Both DMA and NO-DMA increased levels of NOin WT mice. The diminished response in eNOS (−/−) was significantlyattenuated in DMA relative to NO-DMA treated animals. Data show mean andS.E.M. (n=4-12); *=p<0.05, **=p<0.01, ***=p<0.001 using one-way ANOVAwith Bonferroni's post hoc test.

FIG. 5 shows reversal of memory deficit in STPA in WT and eNOS (−/−)mice. Amnestic memory deficit was induced by i.p. injection of eitherscopolamine (1 mg/kg) or L-NAME (50 mg/kg) 30 min prior to training inC57Bl/6 male mice. SERMs (2 mg/kg) were given 20 min prior to trainingand latency was assessed 24 h after training with animals being removedfrom the task if latency >300 s. All SERMs, except F-DMA, restoredscopolamine-induced deficits in STPA in WT animals. AgainstL-NAME-induced deficit, only NO-DMA showed efficacy in reversing memorydeficits. In eNOS (−/−) animals subject to scopolamine-induced amnesia,only NO-DMA showed efficacy. Data show mean and S.E.M. (n=4-10);**=p<0.01, ***=p<0.001 compared to veh. control using one-way ANOVA withDunnett's post hoc test.

FIG. 6 shows effect of SERMs and NO-SERM on intrinsic (PTT) andextrinsic (aPTT) clotting cascades. Blood was collected by cardiacpuncture and PTT and aPTT were determined 1 or 3 h after i.p. drugadministration (2 mg/kg). The anticoagulant activity of DMA was shorterlived than that of NO-DMA and did not persist in the presence of NOSblockade. Data show mean and S.E.M. (n=2-8); *=p<0.05, **=p<0.01,***=p<0.001 using one-way ANOVA with Bonferroni's post hoc test.

FIG. 7 shows G15 has no effect on LTP in C57Bl/6 mice. LTP was measuredafter TBS in the CA1 region of hippocampal sections from 8 month oldmale C57Bl/6 mice. G15 (100 nM) was added 30 min prior to TBS andcontinued throughout. G15 had no effect on LTP, which reached levelsequal to WT background in FIG. 2 for both controls and G15. Data showmean and S.E.M normalized to baseline (n=5-6).

FIG. 8 shows SERM bioavailability in plasma and CNS of WT and eNOS (−/−)mice. Bioavailability was assessed using LC/MS-MS after liquidextraction with internal standard after i.p. injection of 5 mg/kg DMA.DMA shows substantial bioavailability with preferential retention in thehippocampus up to 2 h after administration. Data show mean and S.E.M.(n=4).

FIG. 9 shows ODQ blocks effect of DMA on LTP in 3×Tg mice. LTP wasmeasured in the CA1 region of hippocampal sections from 16 month male3×Tg mice using the method detailed in manuscript, with the exception ofa variable interburst interval during TBS. ODQ (10 nM) was added 30 minprior to TBS and continued throughout. Data show mean and S.E.M.normalized to baseline (n=9-10).

DETAILED DESCRIPTION

In one aspect, the present invention relates to a compound of formula(I)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. X₁ isselected from the group consisting of alkyl, —O—, —N(H)—, —S—, —S(═O)—,and —C(═O)—. A₃ is selected from the group consisting of alkyl, aryl,cycloalkyl, cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selectedfrom the group consisting of hydrogen, halogen, alkyl, cyano,trifluoromethyl, aryl-heteroaryl wherein the aryl is substituted orunsubstituted (e.g., 1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃.R₁, R₂ and R₃ are each independently selected from the group consistingof hydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and—R₄-G₁. R^(x1), R^(y1) and R^(z1), at each occurrence, are independentlyselected from the group consisting of hydrogen and a pharmaceuticallyacceptable cation. R^(a) is alkyl or —OH. R₄, at each occurrence, isindependently selected from the group consisting of a substituentcomprising 1 to 10 carbon atoms, a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen atom, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one C═O group, and a substituent comprising 1 to 10 carbon atomsand optionally containing at least one nitrogen and/or at least one C═Ogroup. G₁, at each occurrence, is independently selected from the groupconsisting of hydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃is —R₄-G₁, and wherein at least one occurrence of G₁ is —ONO₂.

The present invention may also relate to a compound of formula (I),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The present invention may also relate to a compound of formula (I),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. R₁ and R₂ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R₄, at eachoccurrence, is independently selected from the group consisting of asubstituent comprising 1 to 10 carbon atoms, a substituent comprising 1to 10 carbon atoms and optionally containing at least one nitrogen atom,a substituent comprising 1 to 10 carbon atoms and optionally containingat least one C═O group, and a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen and/or at leastone C═O group. R₅ is selected from the group consisting of hydrogen andC₁-C₃-alkyl-G₁. G₁, at each occurrence, is independently selected fromthe group consisting of hydrogen and —ONO₂, wherein at least one of R₁and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁, and wherein at least oneoccurrence of G₁ is —ONO₂. R^(x1), R^(y1) and R^(z1) are eachindependently selected from the group consisting of hydrogen and apharmaceutically acceptable cation. R^(a) is alkyl or —OH.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ib)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. R₁ andR₂ are each independently selected from the group consisting ofhydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R⁴-G₁.R₄, at each occurrence, is independently selected from the groupconsisting of a substituent comprising 1 to 10 carbon atoms, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen atom, a substituent comprising 1 to 10 carbon atomsand optionally containing at least one C═O group, and a substituentcomprising 1 to 10 carbon atoms and optionally containing at least onenitrogen and/or at least one C═O group. R₅ is selected from the groupconsisting of hydrogen and C₁-C₃-alkyl-G₁. G₁, at each occurrence, isindependently selected from the group consisting of hydrogen and —ONO₂,wherein at least one of R₁ and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁,and wherein at least one occurrence of G₁ is —ONO₂. R^(x1), R^(y1) andR^(z1) are each independently selected from the group consisting ofhydrogen and a pharmaceutically acceptable cation. R^(a) is alkyl or—OH.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ic)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R^(x1), R^(y1)and R^(z1), at each occurrence, are independently selected from thegroup consisting of hydrogen and a pharmaceutically acceptable cation.R^(a) is alkyl or —OH. R₄, at each occurrence, is independently selectedfrom the group consisting of a substituent comprising 1 to 10 carbonatoms, a substituent comprising 1 to 10 carbon atoms and optionallycontaining at least one nitrogen atom, a substituent comprising 1 to 10carbon atoms and optionally containing at least one C═O group, and asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen and/or at least one C═O group. G₁, at eachoccurrence, is independently selected from the group consisting ofhydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃ is —R₄-G₁, andwherein at least one occurrence of G₁ is —ONO₂.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄-G₁ is

Representative examples of compounds of formula (I), formula (Ia),formula (Ib), and formula (Ic) may include, but are not limited to:

-   3-(1-(2-(4-((6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-2-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(1-(2-(4-((6-hydroxy-3-(4-hydroxyphenyl)benzo[b]thiophen-2-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-3-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(4-fluorophenyl)-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   or pharmaceutically acceptable salts thereof.

The compounds of formula (I), formula (Ia), formula (Ib), and/or formula(Ic) may be a nitric oxide (NO) donating selective estrogen receptormodulator (NO-SERM). The compounds of formula (I), formula (Ia), formula(Ib), and/or formula (Ic) may prevent or reduce the risk of thrombosisthrough the anticoagulant intrinsic and/or extrinsic pathways. Thecompounds of formula (I), formula (Ia), formula (Ib), and/or formula(Ic) may prevent or reduce the risk of thrombosis even when eNOS isinhibited or inactive. The compounds of formula (I), formula (Ia),formula (Ib), and/or formula (Ic) may be procognitive andneuroprotective even in the presence of NOS dysfunction.

The present invention also relates to methods of treatment ofestrogen-related medical disorders. The methods of treatment may includeadministering to a subject in need of such treatment a compositioncomprising a therapeutically effective amount of at least one compoundof formula (I), formula (Ia), formula (Ib), formula (Ic), and/orpharmaceutically acceptable salts thereof.

An estrogen-related medical disorder may be selected from the groupconsisting of cancer, inflammation, osteoporosis, vaginal atrophy,central nervous system diseases (e.g., Alzheimer's Disease and mildcognitive impairment), and cardiovascular system diseases (e.g.,thrombosis). The cancer may be selected from the group consisting ofbreast cancer, ovarian cancer, prostate cancer, and lung cancer. Thebreast cancer may be a tamoxifen resistant breast cancer or a triplenegative breast cancer.

The present invention also relates to methods of identifying a cancer ina subject. The methods may include obtaining a test sample from thesubject having cancer and determining an amount of PKCα in the testsample. If the amount of PKCα in the test sample is greater than anamount of PKCα in a test sample from a subject not having cancer, thenthe cancer may be sensitive to at least one compound of formula (I),formula (Ia), formula (Ib), formula (Ic), and/or pharmaceuticallyacceptable salts thereof.

In another aspect, the present invention relates to a compound offormula (II),

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), and —C(═O)R^(a). R^(x1), R^(y1) andR^(z1), at each occurrence, are independently selected from the groupconsisting of hydrogen and a pharmaceutically acceptable cation. R^(a)is alkyl or —OH.

Also provided are compounds of formula (II), wherein A₁ is fluoro. A₁may be fluoro located in the 4-position of the phenyl ring to which itis attached. Also provided are compounds of formula (II), wherein A₂ is—OH.

Representative examples of compounds of formula (II) may include, butare not limited to:

-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;-   cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;-   1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;-   1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;-   (4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(p-tolyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   or pharmaceutically acceptable salts thereof.

The compound of formula (II) may be a selective estrogen receptormodulator (SERM). The compounds of formula (II) may prevent or reducethe risk of thrombosis through the anticoagulant intrinsic and/orextrinsic pathways. The compounds of formula may prevent or reduce therisk of thrombosis even when eNOS is inhibited or inactive. Thecompounds of formula (II) may be procognitive and neuroprotective evenin the presence of NOS dysfunction.

The present invention also relates to methods of treatment ofestrogen-related medical disorders. The methods of treatment may includeadministering to a subject in need of such treatment a compositioncomprising a therapeutically effective amount of at least one compoundof formula (II), and/or pharmaceutically acceptable salts thereof.

An estrogen-related medical disorder may be selected from the groupconsisting of cancer, inflammation, osteoporosis, vaginal atrophy,central nervous system diseases (e.g., Alzheimer's Disease and mildcognitive impairment), and cardiovascular system diseases (e.g.,thrombosis). The cancer may be selected from the group consisting ofbreast cancer, ovarian cancer, prostate cancer, and lung cancer. Thebreast cancer may be a tamoxifen resistant breast cancer or a triplenegative breast cancer.

The present invention also relates to methods of identifying a cancer ina subject. The methods may include obtaining a test sample from thesubject having cancer and determining an amount of PKCα in the testsample. If the amount of PKCα in the test sample is greater than anamount of PKCα in a test sample from a subject not having cancer, thenthe cancer may be sensitive to at least one compound of formula (II),and/or pharmaceutically acceptable salts thereof.

1. DEFINITIONS

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art. In case of conflict, the present document, includingdefinitions, will control. Preferred methods and materials are describedbelow, although methods and materials similar or equivalent to thosedescribed herein can be used in practice or testing of the presentinvention. All publications, patent applications, patents and otherreferences mentioned herein are incorporated by reference in theirentirety. The materials, methods, and examples disclosed herein areillustrative only and not intended to be limiting.

The terms “comprise(s),” “include(s),” “having,” “has,” “can,”“contain(s),” and variants thereof, as used herein, are intended to beopen-ended transitional phrases, terms, or words that do not precludethe possibility of additional acts or structures. The singular forms“a,” “and” and “the” include plural references unless the contextclearly dictates otherwise. The present disclosure also contemplatesother embodiments “comprising,” “consisting of” and “consistingessentially of,” the embodiments or elements presented herein, whetherexplicitly set forth or not.

The term “alkoxy” as used herein, refers to an alkyl group, as definedherein, appended to the parent molecular moiety through an oxygen atom.Representative examples of alkoxy include, but are not limited to,methoxy, ethoxy, propoxy, 2-propoxy, butoxy and tert-butoxy.

The term “alkoxycarbonyl” as used herein, refers to an alkoxy group, asdefined herein, appended to the parent molecular moiety through acarbonyl group, as defined herein. Representative examples ofalkoxycarbonyl include, but are not limited to, methoxycarbonyl,ethoxycarbonyl, and tert-butoxycarbonyl.

The term “alkyl” as used herein, refers to a straight or branched chainhydrocarbon containing from 1 to 10 carbon atoms. Representativeexamples of alkyl include, but are not limited to, methyl, ethyl,n-propyl, isopropyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, pentyl,hexyl, heptyl, octyl, nonyl, and decyl. The alkyl groups of thisinvention may be unsubstituted or substituted with 1, 2, or 3 oxosubstituents.

The term “alkylene”, as used herein, refers to a divalent group derivedfrom a straight or branched chain hydrocarbon of 1 to 10 carbon atoms,for example, of 2 to 5 carbon atoms. Representative examples of alkyleneinclude, but are not limited to, —CH₂CH₂—, —CH₂CH₂CH₂—, —CH₂CH₂CH₂CH₂—,and —CH₂CH₂CH₂CH₂CH₂—.

The term “alkylheterocyclealkyl”, as used herein, refers to an alkylgroup, as defined herein, appended to a heterocycle group, as definedherein, wherein the heterocycle group is appended to the parentmolecular moiety through another alkyl group, as defined herein.Representative examples of alkylheteroalkyl include, but are not limitedto, ethylpiperidinylpropyl.

The term “aryl” as used herein, refers to a phenyl group, or a bicyclicfused ring system. Bicyclic fused ring systems are exemplified by aphenyl group appended to the parent molecular moiety and fused to acycloalkyl group, as defined herein, a phenyl group, a heteroaryl group,as defined herein, or a heterocycle, as defined herein. Representativeexamples of aryl include, but are not limited to, indolyl, naphthyl,phenyl, quinolinyl and tetrahydroquinolinyl.

The term “carbonyl” as used herein, refers to a —C(═O)— group.

The term “cycloalkyl” as used herein, refers to a carbocyclic ringsystem containing three to ten carbon atoms, zero heteroatoms and zerodouble bonds. Representative examples of cycloalkyl include, but are notlimited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,cycloheptyl, cyclooctyl, cyclononyl and cyclodecyl. The cycloalkylgroups of this invention may be optionally substituted with 1, 2 or 3alkyl substituents.

The term “cycloalkylalkyl” as used herein, refers to a cycloalkyl group,as defined herein, appended to the parent molecular moiety through analkyl group, as defined herein. Representative examples ofcycloalkylalkyl include, but are not limited to, cyclopropylmethyl,1-methylcyclopropylmethyl, cyclohexylmethyl and3,5-dimethylcyclohexylmethyl.

The term “halogen” as used herein, means Cl, Br, I, or F.

The term “heteroaryl” as used herein, refers to an aromatic monocyclicring or an aromatic bicyclic ring system. The aromatic monocyclic ringsare five or six membered rings containing at least one heteroatomindependently selected from the group consisting of N, O and S. The fivemembered aromatic monocyclic rings have two double bonds and the sixmembered six membered aromatic monocyclic rings have three double bonds.The bicyclic heteroaryl groups are exemplified by a monocyclicheteroaryl ring appended to the parent molecular moiety and fused to amonocyclic cycloalkyl group, as defined herein, a monocyclic aryl group,as defined herein, a monocyclic heteroaryl group, as defined herein, ora monocyclic heterocycle, as defined herein. Representative examples ofheteroaryl include, but are not limited to, indolyl, pyridinyl(including pyridin-2-yl, pyridin-3-yl, pyridin-4-yl), pyrimidinyl andquinolinyl.

The term “heterocycle” as use herein, refers to a non-aromaticmonocyclic ring or a non-aromatic bicyclic ring. The non-aromaticmonocyclic ring is a three, four, five, six, seven, or eight memberedring containing at least one heteroatom, independently selected from thegroup consisting of N, O and S. Representative examples of monocyclicring systems include, but are not limited to, azetidinyl (includingazetidin-2-yl, azetidin-3-yl), piperidinyl (including piperidin-2-yl,piperidin-3-yl, piperidin-4-yl), tetrahydrofuranyl (includingtetrahydrofuran-2-yl, tetrahydrofuran-3-yl) and tetrahydropyranyl(including tetrahydro-2H-pyran-2-yl, tetrahydro-2H-pyran-3-yl,tetrahydro-2H-pyran-4-yl). The bicyclic heterocycles are exemplified bya monocyclic heterocycle appended to the parent molecular moiety andfused to a monocyclic cycloalkyl heterocycle, as defined herein.Bicyclic ring systems are also exemplified by a bridged monocyclic ringsystem in which two non-adjacent atoms of the monocyclic ring are linkedby a bridge of between one and three additional atoms selected from thegroup consisting of carbon, nitrogen and oxygen.

The heterocycles of this invention may be optionally substituted with 1,2 or 3 substituents independently selected from alkyl, alkoxycarbonyl,and oxo.

The term “heterocyclealkyl” as used herein, refers to a heterocyclegroup, as defined herein, appended to the parent molecular moietythrough an alkyl group, as defined herein. Representative examples ofheterocyclealkyl include, but are not limited to ethylpiperidinyl and1-ethylpiperidinyl.

The term “oxo” as used herein, refers to a ═O group.

The term “pharmaceutically acceptable cation” refers to a positivelycharged molecule or atom that is balanced by a negatively chargedmolecule or atom. Representative pharmaceutically acceptable cationsinclude metal salts such as, for example, aluminum, calcium, copper,ferric, ferrous, lithium, magnesium, manganic salts, manganous,potassium, sodium, zinc, and the like as well as nontoxic ammonium,quaternary ammonium and amine cations including, but not limited toammonium, tetramethylammonium, tetraethylammonium, methylamine,dimethylamine, trimethylamine, triethylamine, ethylamine, naturallyoccurring substituted amine, cyclic amines, arginine, betaine, caffeine,choline, N,N′-dibenzylethylenediamine, diethylamine,2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine,ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucamine,glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperidine, procaine, purines,theobromine, tripropylamine, tromethamine, triethanolamine and the like.

The term “trifluoromethyl” as used herein means a —CF₃ group.

In some instances, the number of carbon atoms in a hydrocarbylsubstituent (e.g., alkyl or cycloalkyl) is indicated by the prefix“C_(x)-C_(y)—”, wherein x is the minimum and y is the maximum number ofcarbon atoms in the substituent. Thus, for example, “C₁-C₃-alkyl” refersto an alkyl substituent containing from 1 to 3 carbon atoms.

For the recitation of numeric ranges herein, each intervening numberthere between with the same degree of precision is explicitlycontemplated. For example, for the range of 6-9, the numbers 7 and 8 arecontemplated in addition to 6 and 9, and for the range 6.0-7.0, thenumber 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, and 7.0 areexplicitly contemplated.

2. COMPOUNDS OF THE INVENTION

Compounds of the invention (also referred to herein as “agents”) includecompounds of formula (I)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. X₁ isselected from the group consisting of alkyl, —O—, —N(H)—, —S—, —S(═O)—,and —C(═O)—. A₃ is selected from the group consisting of alkyl, aryl,cycloalkyl, cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selectedfrom the group consisting of hydrogen, halogen, alkyl, cyano,trifluoromethyl, aryl-heteroaryl wherein the aryl is substituted orunsubstituted (e.g., 1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃.R₁, R₂ and R₃ are each independently selected from the group consistingof hydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and—R₄-G₁. R^(x1), R^(y1) and R^(z1), at each occurrence, are independentlyselected from the group consisting of hydrogen and a pharmaceuticallyacceptable cation. R^(a) is alkyl or —OH. R₄, at each occurrence, isindependently selected from the group consisting of a substituentcomprising 1 to 10 carbon atoms, a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen atom, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one C═O group, and a substituent comprising 1 to 10 carbon atomsand optionally containing at least one nitrogen and/or at least one C═Ogroup. G₁, at each occurrence, is independently selected from the groupconsisting of hydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃is —R₄-G₁, and wherein at least one occurrence of G₁ is —ONO₂.

The agent may be a compound of formula (I), wherein R₄, at eachoccurrence, is independently selected from the group consisting ofalkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The agent may be a compound of formula (I), wherein R₄, at eachoccurrence, is independently selected from the group consisting ofalkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. R₁ and R₂ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R₄, at eachoccurrence, is independently selected from the group consisting of asubstituent comprising 1 to 10 carbon atoms, a substituent comprising 1to 10 carbon atoms and optionally containing at least one nitrogen atom,a substituent comprising 1 to 10 carbon atoms and optionally containingat least one C═O group, and a substituent comprising 1 to 10 carbonatoms and optionally containing at least one nitrogen and/or at leastone C═O group. R₅ is selected from the group consisting of hydrogen andC₁-C₃-alkyl-G₁. G₁, at each occurrence, is independently selected fromthe group consisting of hydrogen and —ONO₂, wherein at least one of R₁and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁, and wherein at least oneoccurrence of G₁ is —ONO₂. R^(x1), R^(yl) and R^(z1) are eachindependently selected from the group consisting of hydrogen and apharmaceutically acceptable cation. R^(a) is alkyl or —OH.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ia),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ib)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. R₁ andR₂ are each independently selected from the group consisting ofhydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁.R₄, at each occurrence, is independently selected from the groupconsisting of a substituent comprising 1 to 10 carbon atoms, asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen atom, a substituent comprising 1 to 10 carbon atomsand optionally containing at least one C═O group, and a substituentcomprising 1 to 10 carbon atoms and optionally containing at least onenitrogen and/or at least one C═O group. R₅ is selected from the groupconsisting of hydrogen and C₁-C₃-alkyl-G₁. G₁, at each occurrence, isindependently selected from the group consisting of hydrogen and —ONO₂,wherein at least one of R₁ and R₂ is —R₄-G₁ and/or R₅ is C₁-C₃-alkyl-G₁,and wherein at least one occurrence of G₁ is —ONO₂. R^(x1), R^(y1) andR^(z1) are each independently selected from the group consisting ofhydrogen and a pharmaceutically acceptable cation. R^(a) is alkyl or—OH.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ib),wherein R₄-G₁ is

The compound of formula (I) may comprise a compound of formula (Ic)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), —C(═O)R^(a), and —R₄-G₁. R^(x1), R^(y1)and R^(z1), at each occurrence, are independently selected from thegroup consisting of hydrogen and a pharmaceutically acceptable cation.R^(a) is alkyl or —OH. R₄, at each occurrence, is independently selectedfrom the group consisting of a substituent comprising 1 to 10 carbonatoms, a substituent comprising 1 to 10 carbon atoms and optionallycontaining at least one nitrogen atom, a substituent comprising 1 to 10carbon atoms and optionally containing at least one C═O group, and asubstituent comprising 1 to 10 carbon atoms and optionally containing atleast one nitrogen and/or at least one C═O group. G₁, at eachoccurrence, is independently selected from the group consisting ofhydrogen and —ONO₂, wherein at least one of R₁, R₂ and R₃ is —R₄-G₁, andwherein at least one occurrence of G₁ is —ONO₂.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, cycloalkyl, cycloalkylalkyl, heterocyclealkyl, andalkylheterocyclealkyl, wherein any carbon atom on the alkyl group,together with an alkylene, may form a cycloalkyl, and wherein alkyl maybe unsubstituted or substituted with 1, 2, or 3 oxo substituents.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄, at each occurrence, is independently selected from the groupconsisting of alkyl, alkylheterocyclealkyl, and heterocyclealkyl.

The compound of formula (I) may comprise a compound of formula (Ic),wherein R₄-G₁ or

A compound of formula (I) may include, but may not be limited to

-   3-(1-(2-(4-((6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-2-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(1-(2-(4-((6-hydroxy-3-(4-hydroxyphenyl)benzo[b]thiophen-2-yl)oxy)phenoxy)ethyl)piperidin-4-yl)propyl    nitrate;-   4-(6-hydroxy-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-3-yl)phenyl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   3-(4-fluorophenyl)-2-(4-(2-(piperidin-1-yl)ethoxy)phenoxy)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(trifluoromethyl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(cyclopropanecarbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(cyclopropanecarbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isonicotinoylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-isobutyrylbenzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-ethynylbenzoyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-ethynylbenzoyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-methylbenzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-(4-fluorophenyl)-3-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-(4-fluorophenyl)-2-(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)benzoyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((3-((3r,5r,7r)-adamantane-1-carbonyl)-2-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl    1-((nitrooxy)methyl)cyclopropanecarboxylate;-   (1-(((2-((3r,5r,7r)-adamantane-1-carbonyl)-3-(4-fluorophenyl)benzo[b]thiophen-6-yl)oxy)methyl)cyclopropyl)methyl    nitrate;-   and pharmaceutically acceptable salts thereof.

Compounds of the invention (also referred to herein as “agents”) includecompounds of formula (II)

or a pharmaceutically acceptable salt thereof. A₁ is selected from thegroup consisting of halogen, trifluoromethyl, and —OR₁. A₂ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₂. A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl. A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted (e.g.,1-(4-fluorophenyl)-1H-1,2,3-triazolyl), and —OR₃. R₁, R₂ and R₃ are eachindependently selected from the group consisting of hydrogen, alkyl,—SO₃R^(x1), —PO₃R^(y1)R^(z1), and —C(═O)R^(a). R^(x1), R^(y1) andR^(z1), at each occurrence, are independently selected from the groupconsisting of hydrogen and a pharmaceutically acceptable cation. R^(a)is alkyl or —OH.

Also provided are compounds of formula (II), wherein A₁ is fluoro. A₁may be fluoro located in the 4-position of the phenyl ring to which itis attached. Also provided are compounds of formula (II), wherein A₂ is—OH.

Representative examples of compounds of formula (II) may include, butare not limited to:

-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;-   cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;-   1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;-   1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;-   (4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(p-tolyl)methanone;-   (3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   (3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;-   (3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;-   and pharmaceutically acceptable salts thereof.

Compounds of the invention may act or function as an agonist, anantagonist, a selective estrogen receptor modulator (SERM), or aselective estrogen mimic (SEM). The compounds of may be a nitric oxide(NO) donating selective estrogen receptor modulator (NO-SERM). Thecompounds may prevent or reduce the risk of thrombosis through theanticoagulant intrinsic and/or extrinsic pathways. The compounds of mayprevent or reduce the risk of thrombosis even when eNOS is inhibited orinactive. The compounds may be procognitive and neuroprotective even inthe presence of NOS dysfunction.

The present compounds may exist as a pharmaceutically acceptable salt.The term “pharmaceutically acceptable salt” refers to salts orzwitterions of the compounds which are water or oil-soluble ordispersible, suitable for treatment of disorders without undue toxicity,irritation, and allergic response, commensurate with a reasonablebenefit/risk ratio and effective for their intended use. The salts maybe prepared during the final isolation and purification of the compoundsor separately by reacting an amino group of the compounds with asuitable acid. For example, a compound may be dissolved in a suitablesolvent, such as but not limited to methanol and water and treated withat least one equivalent of an acid, like hydrochloric acid. Theresulting salt may precipitate out and be isolated by filtration anddried under reduced pressure. Alternatively, the solvent and excess acidmay be removed under reduced pressure to provide a salt. Representativesalts include acetate, adipate, alginate, citrate, aspartate, benzoate,benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate,digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate,formate, isethionate, fumarate, lactate, maleate, methanesulfonate,naphthylenesulfonate, nicotinate, oxalate, pamoate, pectinate,persulfate, 3-phenylpropionate, picrate, oxalate, maleate, pivalate,propionate, succinate, tartrate, thrichloroacetate, trifluoroacetate,glutamate, para-toluenesulfonate, undecanoate, hydrochloric,hydrobromic, sulfuric, phosphoric and the like. The amino groups of thecompounds may also be quaternized with alkyl chlorides, bromides andiodides such as methyl, ethyl, propyl, isopropyl, butyl, lauryl,myristyl, stearyl and the like.

Basic addition salts may be prepared during the final isolation andpurification of the present compounds by reaction of a carboxyl groupwith a suitable base such as the hydroxide, carbonate, or bicarbonate ofa metal cation such as lithium, sodium, potassium, calcium, magnesium,or aluminum, or an organic primary, secondary, or tertiary amine.Quaternary amine salts derived from methylamine, dimethylamine,trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine,pyridine, N,N-dimethylaniline, N-methylpiperidine, N-methylmorpholine,dicyclohexylamine, procaine, dibenzylamine, N,N-dibenzylphenethylamine,1-ephenamine and N,N′-dibenzylethylenediamine, ethylenediamine,ethanolamine, diethanolamine, piperidine, piperazine, and the like, arecontemplated as being within the scope of the present invention.

3. PHARMACEUTICAL COMPOSITIONS

Compounds of the invention may be incorporated into pharmaceuticalcompositions suitable for administration to a subject (such as apatient, which may be a human or non-human).

The pharmaceutical compositions may include a “therapeutically effectiveamount” or a “prophylactically effective amount” of the agent. A“therapeutically effective amount” refers to an amount effective, atdosages and for periods of time necessary, to achieve the desiredtherapeutic result. A therapeutically effective amount of thecomposition may be determined by a person skilled in the art and mayvary according to factors such as the disease state, age, sex, andweight of the individual, and the ability of the composition to elicit adesired response in the individual. A therapeutically effective amountis also one in which any toxic or detrimental effects of a compound ofthe invention [e.g., a compound of formula (I), formula (Ia), formula(Ib), formula (Ic), and/or formula (II)] are outweighed by thetherapeutically beneficial effects. A “prophylactically effectiveamount” refers to an amount effective, at dosages and for periods oftime necessary, to achieve the desired prophylactic result. Typically,since a prophylactic dose is used in subjects prior to or at an earlierstage of disease, the prophylactically effective amount will be lessthan the therapeutically effective amount.

For example, a therapeutically effective amount of a compound of formula(I), formula (Ia), formula (Ib), formula (Ic), or formula (II), may beabout 1 mg/kg to about 1000 mg/kg, about 5 mg/kg to about 950 mg/kg,about 10 mg/kg to about 900 mg/kg, about 15 mg/kg to about 850 mg/kg,about 20 mg/kg to about 800 mg/kg, about 25 mg/kg to about 750 mg/kg,about 30 mg/kg to about 700 mg/kg, about 35 mg/kg to about 650 mg/kg,about 40 mg/kg to about 600 mg/kg, about 45 mg/kg to about 550 mg/kg,about 50 mg/kg to about 500 mg/kg, about 55 mg/kg to about 450 mg/kg,about 60 mg/kg to about 400 mg/kg, about 65 mg/kg to about 350 mg/kg,about 70 mg/kg to about 300 mg/kg, about 75 mg/kg to about 250 mg/kg,about 80 mg/kg to about 200 mg/kg, about 85 mg/kg to about 150 mg/kg,and about 90 mg/kg to about 100 mg/kg.

The pharmaceutical compositions may include pharmaceutically acceptablecarriers. The term “pharmaceutically acceptable carrier,” as usedherein, means a non-toxic, inert solid, semi-solid or liquid filler,diluent, encapsulating material or formulation auxiliary of any type.Some examples of materials which can serve as pharmaceuticallyacceptable carriers are sugars such as, but not limited to, lactose,glucose and sucrose; starches such as, but not limited to, corn starchand potato starch; cellulose and its derivatives such as, but notlimited to, sodium carboxymethyl cellulose, ethyl cellulose andcellulose acetate; powdered tragacanth; malt; gelatin; talc; excipientssuch as, but not limited to, cocoa butter and suppository waxes; oilssuch as, but not limited to, peanut oil, cottonseed oil, safflower oil,sesame oil, olive oil, corn oil and soybean oil; glycols; such aspropylene glycol; esters such as, but not limited to, ethyl oleate andethyl laurate; agar; buffering agents such as, but not limited to,magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-freewater; isotonic saline; Ringer's solution; ethyl alcohol, and phosphatebuffer solutions, as well as other non-toxic compatible lubricants suchas, but not limited to, sodium lauryl sulfate and magnesium stearate, aswell as coloring agents, releasing agents, coating agents, sweetening,flavoring and perfuming agents, preservatives and antioxidants can alsobe present in the composition, according to the judgment of theformulator.

4. METHODS OF TREATMENT

The compounds and compositions of the present invention may be used inmethods for treatment of estrogen-related medical disorders. The methodsof treatment may comprise administering to a subject in need of suchtreatment a composition comprising a therapeutically effective amount ofan agent.

a. Estrogen-Related Disorders

The compositions of the present invention may be used in methods fortreatment of estrogen-related medical disorders. An estrogen-relatedmedical disorder may be any medical disorder in which the activity of anestrogen receptor is altered or changed. Alteration of the activity ofan estrogen receptor may include upregulation or downregulation ofestrogen receptor activity. Alteration of the activity of an estrogenreceptor may be the same or different in organs, tissues, and/or cellsof a subject.

An estrogen-related medical disorder may also be any medical disorderresponsive to modulation of the activity of an estrogen receptor. Suchmodulation of the activity of an estrogen receptor may includeupregulation or downregulation of estrogen receptor activity. Theactivity of an estrogen receptor may be modulated or altered by anagonist, an antagonist, a selective estrogen receptor modulator (SERM),a selective estrogen mimic (SEM), a nitric oxide donating SERM(NO-SERM), and/or nitric oxide donating SEM (NO-SEM). The activity ofthe estrogen receptor may be modulated the same or differently indifferent organs, tissues, and/or cells of a subject.

An estrogen-related medical disorder may further be any medical disordercaused by the action of estrogen and/or lack or blocking of estrogenaction. An estrogen-related medical disorder may be any medical disorderresponsive to the action of a composition of the present inventionand/or lack or blocking of estrogen action by a composition of thepresent invention.

An estrogen-related medical disorder may be, but is not limited to,cancer, inflammation, osteoporosis, vaginal atrophy, central nervoussystem diseases, and cardiovascular diseases.

(1) Cancer

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example, cancer.The cancer may be a breast cancer, a uterine cancer, an ovarian cancer,a prostate cancer, and a lung cancer. Particularly, the breast cancermay be a tamoxifen resistant breast cancer or a triple negative breastcancer. The expression or over expression of PKCα may be indicative ofor associated with breast cancer. PKCα may be a biomarker or marker ofbreast cancer.

The method of treatment may prevent or reduce the risk of cancer. Themethod of treatment may cause partial or complete regression of cancerin a subject.

The method of treatment may antagonize estrogen action in the breast.The method of treatment may block or limit the mitogenic activities ofestrogen in the breast, reproductive system, and the prostate. Themethod of treatment may cause partial or complete regression of atamoxifen resistant cancer or tumor. The method of treatment may causepartial or complete regression of a triple negative breast cancer.

(2) Inflammation

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example,inflammation. The method of treatment may prevent or reduce inflammationin a subject in need of such treatment.

(3) Osteoporosis

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example,osteoporosis. The methods of treatment may prevent or reduceosteoporosis in a subject in need of such treatment. The methods oftreatment may prevent or reduce the loss of bone mineral density in asubject. The methods of treatment may reduce or decrease the rate ofbone turnover or fractures. The methods of treatment may improve ormaintain bone mineral density in a subject.

(4) Vaginal Atrophy

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example, vaginalatrophy. The methods of treatment may prevent or reduce vaginal atrophyin a subject in need of such treatment.

(5) Cardiovascular System Diseases

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example,cardiovascular system diseases. The methods of treatment may enhance ormaintain the vasodilatory effects of estradiol on the coronaryvasculature. The methods of treatment may decrease or lower low-densitylipoprotein cholesterol (LDL-C) levels. The methods of treatment mayraise high-density lipoprotein cholesterol (HDL-C) levels. The methodsof treatment may decrease or reduce the risk of myocardial infarction.The methods of treatment may prevent or reduce the risk of thrombosis.The methods of treatment may prevent or reduce the risk of stroke. Themethods of treatment may prevent or reduce the risk of coronary heartdisease.

(6) Central Nervous System Diseases

The methods of the present invention may be used in methods fortreatment of estrogen-related medical disorders, for example, centralnervous system diseases. The disease of the central nervous system maybe Alzheimer's Disease or mild cognitive impairment. The methods oftreatment may prevent, reduce, or reverse Alzheimer's Disease or mildcognitive impairment in a subject in need of such treatment. The methodsof the present invention may reverse cognitive deficits. The methods ofthe present invention may restore cognition, long term potentiation(LTP), and synaptic function, and/or be neuroprotective. The methods ofpresent invention may promote neuronal survival and/or hippocampalneurogenesis.

b. Modes of Administration

Methods of treatment may include any number of modes of administeringthe composition of the present invention. Modes of administration mayinclude tablets, pills, dragees, hard and soft gel capsules, granules,pellets, aqueous, lipid, oily or other solutions, emulsions such asoil-in-water emulsions, liposomes, aqueous or oily suspensions, syrups,elixirs, solid emulsions, solid dispersions or dispersible powders. Forthe preparation of pharmaceutical compositions for oral administration,the agent may be admixed with commonly known and used adjuvants andexcipients such as for example, gum arabic, talcum, starch, sugars (suchas, e.g., mannitose, methyl cellulose, lactose), gelatin, surface-activeagents, magnesium stearate, aqueous or non-aqueous solvents, paraffinderivatives, cross-linking agents, dispersants, emulsifiers, lubricants,conserving agents, flavoring agents (e.g., ethereal oils), solubilityenhancers (e.g., benzyl benzoate or benzyl alcohol) or bioavailabilityenhancers (e.g. Gelucire™). In the pharmaceutical composition, the agentmay also be dispersed in a microparticle, e.g. a nanoparticulatecomposition.

For parenteral administration, the agent can be dissolved or suspendedin a physiologically acceptable diluent, such as, e.g., water, buffer,oils with or without solubilizers, surface-active agents, dispersants oremulsifiers. As oils for example and without limitation, olive oil,peanut oil, cottonseed oil, soybean oil, castor oil and sesame oil maybe used. More generally spoken, for parenteral administration, the agentcan be in the form of an aqueous, lipid, oily or other kind of solutionor suspension or even administered in the form of liposomes ornano-suspensions.

The term “parenterally,” as used herein, refers to modes ofadministration which include intravenous, intramuscular,intraperitoneal, intrasternal, subcutaneous and intraarticular injectionand infusion.

5. METHOD OF IDENTIFICATION

The present invention also relates to methods of identifying a cancer ina subject. The method may include obtaining a test sample from thesubject having cancer and determining an amount of PKCα in the testsample. If the amount of PKCα in the test sample is greater than anamount of PKCα in a test a sample from a subject not having cancer, thenthe cancer is sensitive to at least one compound of the invention [e.g.,a compound of formula (I), formula (Ia), formula (Ib), formula (Ic),and/or formula (II)].

A cancer sensitive to at least one compound of the invention mayoverexpress protein kinase C alpha (PKCα) and/or be resistant totamoxifen. A subject having a cancer sensitive to at least once compoundof the invention may have an amount of PKCα that is greater than asubject not having cancer and/or not having a cancer sensitive to atleast once compound of the invention.

The present invention has multiple aspects, illustrated by the followingnon-limiting examples.

6. EXAMPLES

In the examples below, DMA analogues and NO-DMA were prepared for studyof neuroprotective and procognitive effects of SERMs and of thepotential for an NO-SERM to overcome adverse effects associated withthrombus formation and eNOS dysfunction. We demonstrate that SERMneuroprotection is mediated through a GPR30-dependent mechanism, andextend these results to show that GPR30 mediates the actions of SERMs inrestoration of synaptic transmission in an AD transgenic mouse model. Wealso demonstrate that the procognitive and vasodilatory effects of SERMsare dependent on intact NOS signaling, whereas NO-SERMs preserve actionin models where NOS signaling is impaired. This is the first report ofan NO-SERM that acts as an antithrombotic agent able to reverse deficitsin synaptic transmission and memory in mouse models, while preserving ERbinding and retaining efficacy in the face of attenuated eNOS activity.

Example 1 Materials and Methods for Examples 2-8

Synthesis of NO-DMA (i.e., Compound 8) and DMA Analogues.

Synthesis of F-DMA and DMA has been described (96). Synthesis of NO-DMA(3-(1-(2-(4-(6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yloxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate) and full characterization data are supplied in Example 11.

Animal Use.

Use of animals was approved by the Institutional Animal Care and UseCommittee at the University of Illinois at Chicago. All experimentsconformed to the Animal Welfare Act, Guide to Use and Care of LaboratoryAnimals, and the US Government Principles of the Utilization and Care ofVertebrate Animals Used in Testing, Research and Training guidelines onthe ethical use of animals.

Primary Neuron Preparation.

Primary cultures of dissociated cortical neurons were prepared asfollows; briefly, cortices were dissected from E16-18 Sprague-Dawleyrats (Charles River). After removal of meninges, cortices were manuallydissociated and plated at a density of 10⁶ cells/mL into 96-well plates.Twenty-four hours after plating, cultures were grown in Neurobasal Amedia supplemented with B27, glutamine and pen/strep, with media changesevery 3-4 days. In accordance with published protocols, measured levelsof glia in culture after DIV2 are <0.5%.

Oxygen Glucose Deprivation (OGD).

The cell media was changed to phenol red-free growing media at least 3 hbefore OGD. Receptor blockers were added 45 min prior to the start ofOGD, and SERMs were added at the start of OGD, with concentrationsmaintained through media changes. For OGD, the cells were transferred toa hypoxia chamber controlled at 5% CO₂ and <1% O₂, and growth media wasreplaced with a physiological salt solution (in mM: NaCl 116, CaCl₂ 1.8,MgSO₄ 0.8, KCl 5.4, NaHCO₃ 14.7, NaHPO₄ 1, HEPES 10, pH=7.4). After 2 hOGD, cells were removed from the chamber and growth media was replaced.After 24 h, cell viability was measured by MTT assay using a Dynex MRX11 micro-plate spectrophotometer.

Electrophysiology.

All experiments used 16-month old male 3×Tg transgenic or age-matched WTbackground controls. For electrophysiology, mice were rapidlydecapitated, and brains were removed into an ice-cold aCSF solution (inmM: NaCl 124, KCl 3.0, KH₂PO₄ 1.25, NaHCO₃ 25.7, D-glucose 10,L-ascorbate 2.0, MgSO₄ 2.5, and CaCl₂ 3.3) and sectioned on a tissuechopper into 400 um sections. Slices were transported to a 37° C.solution of aCSF, continuously bubbled with 95% O₂/5% CO₂, and allowedto recover at least 60 min before experimentation. After placement of astimulation electrode in the Schaffer commissural fibers and recordingelectrodes with 2 M NaCl solution into the stratum radiatum of the CA1area, stimulus intensity was set to evoke submaximal fEPSP andcontinuously monitored at 20 s intervals for at least 15 min toestablish a stable baseline. SERMs (100 nM) and G15 (100 nM) wereprepared fresh in perfusate and started at least 30 min before LTPinduction to reach a new baseline after enhancement of basal synaptictransmission was observed (97). LTP was induced using a theta burstinduction protocol by applying 10 bursts of four pulses at 100 Hz withan interburst interval of 200 msec. Resulting fEPSP was monitored at 20s intervals for ˜60 min post TBS.

CNS Bioavailability & NO_(x) Measurement.

All experiments were performed on either male C57Bl/6 mice (CharlesRiver Laboratory) or eNOS (−/−) animals. Animals were injected (i.p.) atvarious time points before sacrifice at the doses specified. Aftereuthanasia via CO₂, plasma was collected, and cortices and hippocampiwere collected after PBS perfusion and stored at −80° C. Tissue sampleswere weighed and extracted with methanol. Plasma samples were extractedwith cold acetonitrile. Quantitative analysis of drug concentrations inplasma and brain used internal standards spiked into plasma and brainhomogenates before liquid extraction with acetonitrile or methanol, andseparation and measurement was performed with LC-MS/MS tandem massspectrometry. The supernatant (deproteinized) was analyzed bychemiluminescence with SIEVERS 280i nitric oxide analyzer.

Step Through Passive Avoidance.

All experiments were performed either on male C57Bl/6 mice or eNOS (−/−)animals. Scopolamine (1 mg/kg) or L-NAME (50 mg/kg) were injected (i.p.)30 min prior to training, while SERMs (2 mg/kg) were injected 20 minprior to training. Mice were placed in the light compartment of thelight/dark box, and as soon as they entered the dark compartment, theyreceived an electric shock (0.5 mA, 60 Hz for 2 seconds). This trainingwas repeated until latency to enter the dark side reached 300 s. At 24 hpost-training, animals were individually placed in the light compartmentand the latency to enter the dark compartment was recorded with a 300 scutoff. Details of this widely used assay of behavior have beendiscussed previously for NO-donor compounds (51, 98, 99).

Aortic Ring Relaxation.

Isolated rings of aortae from male Sprague-Dawley rats (250-300 g) wereprepared for isometric tension measurements and were equilibrated for 1h at an optimal resting tension of 10 mN. Tissues were contractedsubmaximally with 0.3 μM phenylepinephrine, and after 10-15 m cumulativeconcentration-response curves for SERMs (0.01-30 μM) were obtained. Someaortic ring preparations were denuded of the endothelium, or weretreated with 100 μM L-NAME, prior to assessment of SERM-inducedrelaxation responses.

Anticoagulation.

SERMs were injected (i.p.) 1 h before blood collection, and L-NAME wasinjected (i.p.) 30 min before blood collection. The mouse was sedated byexposing it to CO₂ until unconscious and blood was collected in 4.5 mLBD Vacutainer Glass Evacuated Blood Collection Tubes (with 0.105 Mbuffered sodium citrate) by cardiac puncture. The blood samples werecentrifuged at 300 g for 30 min to separate the plasma from the blood,and then 200 uL of plasma was taken from each sample and the PT and aPTTwere measured using an ACL 7000 Coagulation Analyzer. The analyzerinjects 125 μM of PT-fibrinogen (lyophilized rabbit brain calciumthromboplastin with stabilizers, polybrene, buffer and preservatives) to75 μL of plasma sample to measure the PT, and 75 μL of synthAFAX (0.025M)+100 μL of CaCl₂ (0.02 M) were added to 75 μL of plasma to measureaPTT. The analysis was at 37° C. using a photo sensor at λ=671 nm.

Example 2 Synthesis of an NO-SERM Retaining Nanomolar Binding Affinityfor ER

Structural modification of DMA to introduce NO-donor properties, asdetailed below, was accomplished without loss of ER binding. Ligandbinding affinity was measured by competitive E₂ displacement aspreviously described (52). The IC₅₀ values for ERα and ERPβ measured forNO-DMA were 21.4±6.7 nM and 24.4±9.6 nM, respectively. For comparison,ERα IC₅₀ values of 21.0±2.7 nM, 7.8±1.9 nM, and 17±0.6 nM were measuredfor raloxifene, DMA, and F-DMA, respectively.

Example 3 Neuroprotection is Mediated Through GPR30 Receptor Activation,Independent of ERα and NOS

Previously, we have shown that the neuroprotective actions of raloxifeneand DMA are GPR30-dependent and mediated via PI3K/Akt signaling (53).The oxygen glucose deprivation (OGD) assay was used as a composite modelin primary neuronal culture of ischemia-reperfusion injury to determineif addition of NO-donor capacity to DMA altered the mechanism of action.Twenty-four hours after a 2 h glucose deprivation period, 100 nM NO-DMAwas observed to elicit robust neuroprotection identical to DMA, asmeasured by MTT (FIG. 1) or LDH (data not shown) and normalized toestradiol (E2, 10 nM). Blockade of classical ERα signaling by ICI 182780(100 nM) did not block this effect. However, both pertussis toxin (100ng/mL), a GPR blocker, and G15 (100 nM), a selective GPR30 antagonist(54), blocked the neuroprotective activity of NO-DMA and DMA. LY294002(10 μM), a selective PI3K inhibitor, was effective, whereas L-NAME (100μM), a non-selective NOS antagonist, was ineffective in attenuatingneuroprotective activity, supporting the hypothesis that NO-DMA signalsthrough the PI3K/Akt pathway downstream of GPR30 in a manner similar toDMA. In this experimental model, signaling via NOS downstream of PI3/Aktis not indicated.

Example 4 LTP Restoration in an Alzheimer's Mouse Model isGPR30-Dependent

Long-term potentiation (LTP) in the CA1 field of the hippocampus is awell-studied cellular model for learning and memory. To measure theeffect of DMA and NO-DMA in reversing deficits in synaptic plasticity,LTP was induced with a theta burst stimulation (TBS) protocol atSchaffer/commissural fiber synapses in the CA1 field of hippocampalslices from 16-month old male 3×Tg mice. These mice have been shown tohave a marked deficit in LTP that becomes apparent at 6 months of age(55). At 16 months, an advanced age at which Alzheimer's-likeneuropathology is well developed in these animals, studies on LTP havenot previously been reported. We observed a robust, reproducible deficitin LTP in these aged transgenic mice, with field excitatorypost-synaptic potentials (fEPSP) showing an end average potentiation of97.0±6.4% of baseline at 45 min post-TBS, compared to 134.7±10.3%observed in the wild type (WT) controls (FIG. 2A,B). Addition of 100 nMDMA to the aCSF perfusate 30 min prior to induction of LTP resulted insignificant reversal of the LTP deficit to an end average potentiationof 144.9±8.7% (FIG. 2A,B). NO-DMA (100 nM) had effects similar to DMA,with an average potentiation of 144.2±5.2% of baseline, with a trendtowards increased potentiation over untreated transgenics seenimmediately after TBS (FIG. 2A,B). Finally, addition of the GPR30selective blocker G15 (100 nM) prevented the action of DMA, with endfEPSP approaching levels of untreated transgenics (96.8±7.10%), whilehaving no effect on WT C57Bl/6 slices in the absence of DMA (FIG. 7),suggesting that the GPR30 receptor is critical for the LTP-enhancingeffects of SERMs. Preliminary data also indicates that DMA activity isNOS-dependent (FIG. 9).

To determine if drugs enhanced LTP by altering physiological responsesduring LTP induction, such as NMDA receptor-mediated currents ordownstream events involved in expression or stabilization, thepostsynaptic responses to theta bursts were quantified as previouslydescribed (56, 57). The enhancement of subsequent bursts was augmentedby DMA and NO-DMA but not by DMA+G15 (FIG. 2C), suggesting that SERMsmay act, at least in part, by enhancing depolarization and NMDA receptoractivity during TBS, in addition to downstream actions on the signalingevents that lead to stabilization of the LTP response.

Example 5 eNOS-Dependent Vascular Relaxation is Retained by NO-SERM inthe Absence of eNOS

The vasodilator activity of SERMs and NO-SERMs was assessed usingisolated aortic ring preparations. All SERMs exhibited dose-dependentrelaxation, with raloxifene and DMA showing efficacy approaching 100%relaxation (FIG. 3A). The following EC₅₀ values (mean, S.D.) werecalculated but not found to be significantly different: FDMA, 2.4±1.2μM; arzoxifene, 3.3±2.5 μM; DMA, 3.5±2.5 μM; raloxifene, 4.0±2.9 μM,suggesting equal potency as vasodilators for all SERMs studied. Removalof the endothelium or inhibition of NOS by L-NAME resulted in asignificant decrease in the efficacy of DMA, indicating that themajority of the vasodilator activity of DMA is mediated throughactivation of eNOS (FIG. 3B). Thus, SERMs induce relaxation in theaortic ring model at low micromolar concentrations, dependent on intacteNOS/NO signaling. NO-DMA induced maximal relaxation in this model, withat least ten-fold increased potency (EC₅₀=130±90 nM) over DMA (FIG. 3C).The potency of NO-DMA was reduced in endothelium-denuded tissues (EC₅₀value of 0.90±0.69 μM), and tissues treated with L-NAME (EC₅₀ value of1.0±0.54 μM). NO-DMA was shown to have both endothelium-dependent and-independent vasodilatatory actions, mediated through eNOS and direct NOrelease.

Example 6 CNS Bioavailable SERMs Increase NO Levels Dependent Upon eNOSActivity

The brain and plasma bioavailability of DMA (5 mg/kg; i.p.) was measuredin C57Bl/6 mice using LC/MS-MS detection. DMA showed substantial bloodbrain barrier permeability, with preferential retention in thehippocampus over the cortex (FIG. 8). Brain and plasma NO levels wereassessed by measuring metabolic oxidation products, NO₂ ⁻ and NO₃ ⁻(NO_(x)), using chemiluminescence detection. One hour after injection of2 mg/kg DMA or NO-DMA, there was a greater than twofold elevation ofNO_(x) in the hippocampus and plasma (FIG. 4). To assess the relativecontribution of eNOS, DMA and NO-DMA were administered to eNOS (−/−)animals. In these animals, DMA had no effect on NO_(x) in plasma orhippocampus, whereas NO-DMA was found to increase NO_(x) significantly,in both hippocampus and plasma.

Example 7 SERMs Reverse Cholinergic Cognitive Deficits; NO-SERM Reversesthe Deficit in eNOS KO Mice

To investigate the procognitive effects of SERMs and NO-DMA in an invivo behavioral model of memory, step-through passive avoidance (STPA)was used in C57Bl/6 mice treated with scopolamine (1 mg/kg) 30 min priorto training. In this assay, memory is tested 24 h and 48 h aftertraining and since drugs are only administered during training, any sideeffects of drugs, such as sedation are highly unlikely to confound thetesting results. A high latency reflects a strong consolidated memory ofthe aversive stimulus. In scopolamine treated animals, all treatments (2mg/kg, i.p., 20 min prior to training) except F-DMA and inorganicnitrate induced complete reversal of the memory deficit at 24 h aftertraining (FIG. 5). NO-DMA showed equivalent restoration of protection toDMA. To isolate the procognitive effects due to NO release from NO-DMA,experiments were repeated after administration of L-NAME (50 mg/kg, 30min prior to training) in place of scopolamine. L-NAME alone resulted ina cognitive deficit that was reversed only by NO-DMA. Similar resultswere observed in STPA after a scopolamine-induced deficit in eNOS KOmice, strongly implicating eNOS as mediating the procognitive mechanismof action of SERMs and demonstrating the ability of an NO-SERM tocircumvent loss of eNOS activity.

Example 8 SERMs Act Through the Intrinsic Pathway; NO-SERM ActivatesBoth Intrinsic and Extrinsic Anticoagulant Pathways

NO has been implicated in directly reducing clotting (28), thereforeprothrombin (PT) and activated thromboplastin times (aPTT) wereevaluated for DMA and NO-DMA. At 1 h, both DMA and NO-DMA significantlyincreased aPTT, and both increased PT, but only NO-DMA reachedsignificance (FIG. 6). After 3 h, the effect of DMA had decreased tothat of control samples, whereas the effect of NO-DMA remainedsignificantly higher than control. To test the anticoagulant activity ofcompounds in models of impaired NOS function, mice injected with L-NAME(50 mg/kg, 30 m prior to collection) were administered DMA or NO-DMA (2mg/kg, 1 h prior to collection), and only NO-DMA showed significantanticoagulation compared to control, with the increase in PTsignificantly higher than DMA.

Example 9 Summary of Examples 2-8

Although 2^(nd) generation selective estrogen receptor modulators(SERMs) have overcome gynecological issues, the potential of estrogensin neuroprotective and procognitive therapy is limited by the risk ofthrombotic events. The critical period hypothesis also posits anattenuated response to estrogens with age. The 3^(rd) generation SERM,desmethylarzoxifene (DMA), was shown in hippocampal slices from older3×Tg Alzheimer's transgenic mice to restore synaptic function withdependence on GPR30. Neuroprotection in primary rat neurons was alsoGPR30 dependent. DMA was brain bioavailable and reversed cholinergiccognitive deficits, however, DMA was not able to restore cognition inmice in which endothelial nitric oxide synthase (eNOS) was inhibited ordeleted. A novel NO-donating SERM (NO-SERM) was designed and shown to beprocognitive and neuroprotective even with NOS dysfunction: the NO-SERMrestored cognition in eNOS (−/−) mice, restored LTP in 3×Tg hippocampalslices, and protected primary neurons against ischemia-reperfusioninjury. The activation of eNOS underlies the cardiovascular benefits ofestrogens and relaxation of aortic rings was induced by SERMs, althoughonly NO-DMA retained efficacy in the absence of eNOS. Finally, incontrast to DMA, NO-DMA was shown to reduce thrombosis through both theintrinsic and extrinsic pathway, even when eNOS was inhibited. TheNO-SERM retains the positive attributes of SERMs, without the thromboticside effect, and can be of use in an aging population in which eNOSactivity is attenuated.

Example 10 Discussion of Examples 2-8

Menopause has been associated with dementia, depression, and anxiety,and the management of such effects with safe and effective hormonetherapies is a largely unmet need. Women entering menopause reportmemory and concentration difficulties (58, 59), and the age-adjusted ADmortality rate was shown to be 21.7% higher for women than for men (60).Estrogen replacement therapy has been reported to improve or maintainlevels of cognitive function in postmenopausal women, and to reduce therisk of Alzheimer's disease (AD), observations in accord with proposedroles for estrogen in the brain in protection, growth anddifferentiation of neurons (7, 61-63). However, adverse eventsassociated with estrogen commend SERMs as an alternative approach. Thebeneficial effects of estrogen in the CNS, bone, and cardiovascularsystems underlie the search for an ideal SERM that can preserve thesebeneficial effects while minimizing or antagonizing the potentiallyserious adverse effects of estrogens in sex organs.

The benzothiophene SERM, raloxifene, has demonstrated significantbeneficial effects on bone and lipid profiles in postmenopausal women(17-19), with evidence suggesting that CNS and cardiac profiles remainpositive (14, 23). Furthermore, raloxifene shows antiestrogenic activityin the breast and endometrium (20, 21). Clinical studies on raloxifenehave shown positive signals on cognitive skills (18, 64-66), andprocognitive and neuroprotective effects have been reported in femalerats (67-69). However, raloxifene has also shown efficacy in male rats(70) and positive cognitive responses have been reported in elderly men(71, 72). The benzothiophene SERMs, raloxifene and DMA, have manycharacteristics of an ideal SERM of potential use in the CNS, inparticular if thrombotic side effects can be abolished.

Relevant to design of an ideal SERM is the interplay between classicalER-mediated signaling and emerging mechanisms of rapid extranuclearsignaling, such as via GPR30 (54, 73, 74). Associated with thesesignaling pathways, are important roles for NO in signaling viaactivation of eNOS through the PI3K/Akt pathway (25, 26). Estrogen andSERM induced neuroprotection has been reported to be mediated via rapidsignaling, including regulation of intracellular Ca²⁺ andphosphorylation of ERK (75, 76). Extranuclear ER, linked to kinasesignaling pathways, including PI3K/Akt and Src/ERK/CREB, has been shownto play a role in estrogenic neuroprotection (75, 77), providing directlinks to anti-apoptotic mechanisms (78). Estradiol and DMA were observedto elicit neuroprotection in primary neurons subject toischemia-reperfusion injury, with the novel observation that the actionsof estrogen and SERMs were mediated via GPR30 linked to downstreamPI3K/Akt, Src, and ERK pathways. This activity was retained by NO-DMA, aNO-SERM that is itself a potent antiestrogen; although neuroprotectionin this model system was independent of NOS.

To extend observations on neuronal rescue by SERMs and NO-SERM to amodel system for synaptic rescue, the 3×Tg Alzheimer's mouse model wasused. These mice represent a more complete model of the disease thansome other strains, with progressive production of Aβ in relevant brainareas, plaque and tangle pathology similar to those observed in ADpatients, and synaptic transmission and LTP impairment at 6 months ofage (55, 79). We chose to study synaptic function in 3×Tg mice at 16months, representing a model of aging, combined with substantial Aβ andtau neuropathology. Gratifyingly, DMA was observed to restore synapticdeficits, measured by LTP in 3×Tg hippocampal slices; and this activitywas again dependent upon GPR30 and retained by NO-DMA.

Relaxation of blood vessels by estrogens and SERMs is mediated byendothelium-independent and -dependent mechanisms, the latter involvingactivation of eNOS. ER isoforms and GPR30 have been implicated in theseactions (80). The vascular effects of raloxifene have been wellestablished in conduit arteries, and recently in resistance arteries(81), supporting therapeutic benefit in preserving endothelial function.Raloxifene was first reported to induce vasodilation by an ER andeNOS-dependent pathway (82), and subsequently shown to activate eNOS viaPI3K/Akt (26). In rat aorta, vasodilation was observed in response toall benzothiophene SERMs tested. Relaxation induced by DMA was largelyendothelium and eNOS-dependent, whereas NO-DMA retained equivalentpotency to DMA even in the absence of endothelium and eNOS.

Cholinergic deficits are the basis of current symptomatic therapy of AD(83). The roles that cholinergic dysfunction may play in early AD andMild Cognitive Impairment (MCI) have recently been reviewed (84).Amnestic MCI is a putative prodromal stage of Alzheimer's disease andchemical amnesia has frequently been induced in man and animal models indrug discovery and development using cholinergic blockade withscopolamine (85). Benzothiophene SERMs, raloxifene, arzoxifene, and DMArestored the cognitive deficit induced in mice by scopolamine. Theconcerted actions of eNOS and nNOS are required for normal LTP in thehippocampus, and in tissues from knockout mice, exogenous NO can replaceNOS function (38). Therefore, given observations on SERMs and NO-SERM inthe vasculature, the use of both NOS inhibition and eNOS (KO) mice wasindicated to test the efficacy of DMA and NO-DMA. The procognitivemechanism of action of DMA was identified as being eNOS-dependent;NO-DMA was observed to retain procognitive actions in both paradigms ofeNOS-challenge.

The known endothelial dysfunction associated with aging, the decreasedexpression of eNOS in postmenopausal women, and the potentialcontribution of eNOS to the critical period hypothesis, commend thefurther study of therapeutic approaches that reinforce NO/cGMP signalingin combination with ER modulation. Other contributions to age-relatedpathophysiology, in addition to attenuated eNOS expression, have beenproposed, including: inhibition of eNOS by elevation of ADMA (86, 87);depletion of cofactors, notably BH4(88-90); and eNOS polymorphisms (91).Attenuation of NO-dependent cGMP production by soluble guanylyl cyclase(sGC) has also been linked directly with Aβ. (92) The NO-SERM approachis informative, however, related approaches to combinatorial activationof (1) estrogenic signaling and (2) NO/cGMP signaling are also endorsed(27, 93).

In summary, a prototype NO-SERM was designed and demonstrated to retainthe ER binding and classical ERα antagonist actions of the parent SERM,and to provide neuroprotection and synaptic restoration in an AD model,dependent upon GPR30. Against a cholinergic cognitive deficit in mice,neuroprotective SERMs restored memory, however, this procognitive effectwas lost when eNOS activity was blocked or absent. The NO-SERM restoredcognition after cholinergic insult in eNOS (−/−) mice and NO levelsmeasured after SERM and NO-SERM treatment were compatible with thecentral role of NO signaling. In AD in particular, a role is emergingfor NO in treating cholinergic deficits (49) and attenuatingamyloidogenesis (94, 95).

From a therapeutic perspective, the known endothelial dysfunctionassociated with aging, the decreased expression of eNOS inpostmenopausal women, and the potential contribution of eNOS to thecritical period hypothesis, commend the further study of NO-SERMs.Related approaches to combinatorial activation of (1) estrogenicsignaling and (2) NO/cGMP signaling are also endorsed (27, 89). Theability of an NO-SERM to reduce coagulation by increasing both PT andaPTT to levels comparable to those of clinical anticoagulants, suggeststhat this approach can also address the risk of thromobosis associatedwith SERM therapeutics. These findings, coupled with the otherwiseexcellent safety record of raloxifene in the clinic, support furtherdevelopment of NO-SERMs or combination therapies for AD and otherneurodegenerative disorders in the post-menopausal cohort, and suggestutility for this approach in the general population.

Example 11 Synthesis of NO-DMA Compound 8 (i.e.,3-(1-(2-(4-(6-hydroxy-2-(4-hydroxyphenyl)benzo[b]thiophen-3-yloxy)phenoxy)ethyl)piperidin-4-yl)propylnitrate)

¹H and ¹³C NMR spectra were obtained with Bruker Ultrashield 400 MHz andAdvance 300 MHz spectrometers. Chemical shifts are reported as δ valuesin parts per million (ppm) relative to tetramethylsilane (TMS) for allrecorded NMR spectra. All reagents and solvents were obtainedcommercially from Acros, Aldrich, and Fluka and were used withoutpurification.

Starting material 1 was synthesized from commercial available6-methoxy-2-(4-methoxy phenyl)benzo[b]thiophene according to a publishedmethod in 6 steps (53). Demethylation of 4 gave complex mixture, mightbe due to the incompatibility of Lewis acid with organic nitrate. NO-DMA8 was obtained by the reaction of 5 with silver nitrate in low yield(method A); or by using tetrabutyl ammonium nitrate as nitrationreagent, the yield was improved dramatically, but the method sufferedfrom repetitive column purifications to remove ammonium salt (method B);alternatively, the phenol groups of 5 were acetylated and the bromide 6was converted nitrate 7 using silver salt, 8 was finally obtained withgood purity by de-acetylation of 7 in high yield (method C).

Compound 2.

Compound 1 (2.6 g, 5.3 mmol), 4-piperidinepropanol (1.0 g, 6.9 mmol)were dissolved in anhydrous acetonitrile (100 mL) and anhydrous K₂CO₃(2.9 g, 21.5 mmol) was added. The reaction mixture was refluxed for 6hrs and cooled to room temperature. After filtration, solvent wasremoved, the residue was dissolved in DCM (150 mL) and washed with water(2×50 mL). Organic phase separated and concentrated. The residue waspurified by column chromatography (DCM/MeOH 20:1, 2.9 g, quantitativeyield). ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.66-7.69 (m, 2H), 7.42 (d, 1H,J=2.0 Hz), 7.24 (d, 1H, J=8.8 Hz), 6.81-6.93 (m, 7H), 3.97 (t, 2H, J=6.0Hz), 3.84 (s, 3H), 3.76 (s, 3H), 3.51 (t, 2H, J=6.4 Hz), 2.88-2.91 (m,2H), 2.63 (t, 2H, J=6.0 Hz), 1.95-2.01 (m, 2H), 1.60-1.62 (m, 2H),1.46-1.53 (m, 2H), 1.21-1.27 (m, 5H); ¹³C-NMR (Acetone-d₆, 100 MHz):160.23, 159.01, 155.19, 152.34, 140.57, 137.56, 129.32, 128.77, 127.06,125.78, 122.64, 116.95, 116.26, 115.28, 115.05, 106.36, 67.30, 62.64,58.28, 55.90, 55.52, 55.13, 36.32, 33.62, 33.31, 30.90.

Compound 3.

Compound 2 (1.1 g, 2.0 mmol) was dissolved in chloroform (25 mL), CBr₄(2.7 g, 8.0 mmol) and polymer bound Ph₃P (2.6 g, about 3 mmol/g) wereadded, the reaction mixture was stirred at r.t. overnight. Afterfiltration, solvent was removed, and the residue was purified by columnchromatography (DCM/AcOEt/MeOH 3:1:0.2, 1.1 g, 89%). ¹H-NMR (Acetone-d₆,400 MHz): 7.68-7.72 (m, 2H), 7.46 (d, 1H, J=2.0 Hz), 7.25 (d, 1H, J=8.8Hz), 6.84-6.97 (m, 7H), 4.00 (t, 2H, J=6.0 Hz), 3.87 (s, 3H), 3.80 (s,3H), 3.47 (s, 2H, J=6.8 Hz), 2.91-2.94 (m, 2H), 2.67 (t, 2H, J=6.0 Hz),1.99-2.04 (m, 2H), 1.82-1.89 (m, 2H), 1.62-1.65 (m, 2H), 1.32-1.37 (m,2H), 1.15-1.24 (m, 3H); δ ¹³C-NMR (Acetone-d₆, 100 MHz): 160.35, 159.12,155.29, 152.43, 140.64, 137.63, 129.38, 128.82, 127.10, 125.63, 122.68,117.00, 116.36, 115.37, 115.12, 106.43, 67.43, 58.30, 55.96, 55.58,55.06, 35.83, 35.78, 34.98, 33.22, 30.98.

Compound 4.

Compound 3 (110 mg, 0.18 mmol) was dissolved in anhydrous acetonitrile(3 mL), AgNO₃ (306 mg, 1.8 mmol) was added. The reaction mixture wasstirred at r.t. for 24 h. Solvent was removed and the residue waspurified by column chromatography (DCM/AcOEt/MeOH 2:1:0.25, 85 mg, 79%).¹H-NMR (Acetone-d₆, 400 MHz): δ 7.66-7.72 (m, 2H), 7.43 (d, 1H, J=2.0Hz), 7.25 (d, 1H, J=8.8 Hz), 6.82-6.95 (m, 7H), 4.49 (t, 2H, J=6.8 Hz),3.98 (t, 2H, J=6.0 Hz), 3.85 (s, 3H), 3.77 (s, 3H), 2.89-2.92 (m, 2H),2.64 (t, 2H, J=2.0 Hz), 1.97-2.02 (m, 2H), 1.67-1.73 (m, 2H), 1.61-1.64(m, 2H), 1.13-1.34 (m, 5H); ¹³C-NMR (Acetone-d₆, 100 MHz): δ 160.30,159.07, 155.25, 152.39, 140.62, 137.61, 129.36, 128.81, 127.09, 125.62,122.67, 116.99, 116.32, 115.32, 115.09, 106.41, 74.69, 67.40, 58.27,55.94, 55.56, 54.99, 36.03, 33.09, 24.64.

Compound 5.

Compound 3 (240 mg, 0.39 mmol) was dissolved in DCM (5 mL), HCl ethersolution (2.0M, 1.0 mL) was added. The reaction mixture was stirred at4° C. for 2 hr. Solvent was removed, the residue was re-dissolved in DCM(10 mL), cooled in an ice-water bath, BBr₃ in DCM (1.0 M, 1 mL) wasadded dropwise. The reaction mixture was stirred at 4° C. for 3 hr andthen at r.t. for another hour. The reaction mixture was poured into icewater (50 mL), neutralized with NaHCO₃ and extracted with AcOEt (3×30mL). Organic solution was combined and concentrated, the residue waspurified by column chromatography (DCM/MeOH 50:3, 160 mg, 70%). ¹H-NMR(Acetone-d₆, 400 MHz): δ 7.59-7.62 (m, 2H), 7.32 (d, 1H, J=2.0 Hz), 7.19(d, 1H, J=8.8 Hz), 6.83-6.89 (m, 7H), 4.06 (t, 2H, J=5.9 Hz), 3.47 (t,2H, J=6.7 Hz), 3.01-3.04 (m, 2H), 2.77 (t, 2H, J=5.9 Hz), 2.11-2.17 (m,2H), 1.82-1.90 (m, 2H), 1.65-1.67 (m, 2H), 1.22-1.38 (m, 5H); ¹³C-NMR(Acetone-d₆, 100 MHz): δ 158.14, 156.63, 154.99, 152.52, 140.31, 137.46,129.41, 128.06, 126.60, 124.59, 122.67, 116.97, 116.54, 116.30, 115.64,108.79, 66.85, 58.04, 54.83, 35.61, 35.52, 34.95, 32.68, 30.90.

Compound 6.

Compound 5 (300 mg, 0.52 mmol) was dissolved in DCM (10 mL), pyridine(0.84 mL, 10.3 mmol) and Ac₂O (0.34 mL, 3.6 mmol) were added, thereaction mixture was stirred at r.t. overnight. Solvent was removed, theresidue was re-dissolved in ethyl acetate (50 mL) and washed with water,organic solution was separated and concentrated, the residue waspurified by column chromatography, product was obtained as slightlyyellow foam (DCM/MeOH 30:1, 298 mg, 87%). ¹H-NMR (Acetone-d₆, 400 MHz):δ 7.83 (d, 2H, J=8.4 Hz), 7.74 (d, 1H, J=2.0 Hz), 7.40 (d, 1H, J=8.8Hz), 7.19 (d, 2H, J=8.8 Hz), 7.10 (dd, 1H, J=8.8 Hz, 2.0 Hz), 6.86-6.93(m, 4H), 4.05 (t, 2H, J=6.0 Hz), 3.47 (t, 2H, J=6.8 Hz), 2.95-2.98 (m,2H), 2.71 (t, 2H, J=6.0 Hz), 2.29 (s, 3H), 2.26 (s, 3H), 2.06-2.11 (m,2H), 1.82-1.89 (m, 2H), 1.64-1.66 (m, 2H), 1.32-1.38 (m, 2H), 1.20-1.28(m, 3H); ¹³C-NMR (Acetone-d₆, 100 MHz): 169.75, 169.46, 155.42, 152.20,151.79, 149.90, 141.64, 136.84, 132.49, 130.24, 129.33, 129.24, 123.25,122.81, 120.61, 117.13, 116.89, 116.46, 67.17, 58.12, 54.93, 35.69,35.66, 34.98, 32.96, 30.94, 20.93.

Compound 7.

Compound 6 (280 mg, 0.45 mmol) was dissolved in anhydrous acetonitrile(15 mL), AgNO₃ (1.1 g, 1.8 mmol) was added. The reaction mixture wasstirred at r.t. for 24 h. After filtration, solvent was removed and theresidue was purified by column chromatography, product was obtained asslightly yellow foam (DCM/MeOH 30:1, 190 mg, 70%). ¹H-NMR (Acetone-d₆,400 MHz): δ ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.81 (d, 2H, J=8.8 Hz), 7.71(d, 1H, J=2.0 Hz), 7.38 (d, 1H, J=8.8 Hz), 7.17 (d, 2H, J=8.8 Hz), 7.08(dd, 1H, J=8.8 Hz, 2.0 Hz), 6.82-6.92 (m, 4H), 4.48 (t, 2H, J=6.8 Hz),4.00 (t, 2H, J=6.0 Hz), 2.92-2.95 (m, 2H), 2.68 (t, 2H, J=6.0 Hz), 2.27(s, 3H), 2.24 (s, 3H), 2.02-2.04 (m, 2H), 1.61-1.74 (m, 4H), 1.16-1.31(m, 5H); ¹³C-NMR (Acetone-d₆, 100 MHz): δ 169.69, 169.41, 155.34,152.12, 151.70, 149.82, 141.58, 136.77, 132.44, 130.19, 129.25, 129.19,123.18, 122.77, 120.53, 117.08, 116.82, 116.38, 74.65, 67.09, 58.07,54.85, 35.83, 32.96, 32.79, 24.58, 20.93.

Compound 8.

Method A: Compound 5 (55 mg, 0.095 mmol) was dissolved in anhydrousacetonitrile (2 mL), AgNO₃ (160 mg, 0.95 mmol) was added. The reactionmixture was stirred at r.t. overnight. After filtration, solvent wasremoved and the residue was purified by column chromatography(DCM/AcOEt/MeOH 3:1:0.3), product was obtained as white foam (15 mg,28%). Method B: Compound 5 (100 mg, 0.17 mmol) was dissolved inanhydrous acetonitrile (5 mL), tetrabutylammonium nitrate (523 mg, 1.7mmol) was added. The reaction mixture was heated at 70° C. for 8 hrs.Solvent was removed and the residue was purified repeatedly by columnchromatography (DCM/AcOEt/MeOH 3:1:0.3) three times, product wasobtained as white foam (68 mg, 70%). Method C: Compound 7 (130 mg, 0.2mmol) was dissolved in a mixture of THF-MeOH—H₂O (4.5 mL, 2:2:0.5),K₂CO₃ (79 mg, 0.58 mmol) was added. The reaction mixture was stirred atr.t. for 1.5 hr. After filtration and concentration, the residue waspurified by column chromatography (DCM/MeOH 20:1), product was obtainedas white foam (103 mg, 91%). ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.60 (d,2H, J=8.8 Hz), 7.31 (d, 1H, J=2.0 Hz), 7.19 (d, 1H, J=8.8 Hz), 6.84-6.90(m, 7H), 4.53 (t, 2H, J=6.7 Hz), 4.03 (t, 2H, J=5.9 Hz), 2.95-2.98 (m,2H), 2.70 (t, 2H, J=5.9 Hz), 1.98-2.04 (m, 2H), 1.73-1.77 (m, 2H),1.66-1.69 (m, 2H), 1.19-1.35 (m, 5H); ¹³C-NMR (Acetone-d₆, 100 MHz): δ158.14, 156.62, 155.11, 152.50, 140.36, 137.49, 129.43, 128.12, 126.63,124.64, 122.71, 116.98, 116.56, 116.29, 115.47, 108.81, 74.69, 67.12,58.21, 54.93, 35.94, 33.03, 32.86, 24.63.

Example 12 Synthesis of NO-SERM 13 and 14

Compound 9.

4′Br-DMA (75 mg, 0.14 mmol) was dissolved in a mixture THF (4 mL) andDIPEA (347 μL, 2.1 mmol), MOM-Br (16 μL, 0.17 mmol) was added. Thereaction mixture was refluxed for 5 hrs. Most of the solvent was removedunder reduced pressure. The residue was dissolved in DCM and washed withaqueous NaHCO₃ and brine. Organic phase separated and concentrated. Theresidue was purified by column chromatography (DCM/MeOH 20:1), productwas obtained as oil (55 mg, 68%). ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.70(d, 2H, J=8.8 Hz), 7.58 (d, 1H, J=2.0 Hz), 7.55 (d, 2H, J=8.4 Hz), 7.30(d, 1H, J=8.8 Hz), 7.02 (dd, 1H, J=8.8 Hz, 2.0 Hz), 6.83-6.90 (m, 4H),5.26 (s, 2H), 3.99 (t, 2H, J=6.0 Hz), 3.45 (s, 3H), 2.64 (t, 2H), 2.44(broad peak, 4H), 1.48-1.54 (m, 4H), 1.38-1.41 (m, 2H); ¹³C-NMR(Acetone-d₆, 100 MHz): 155.89, 155.43, 152.08, 142.33, 137.78, 132.76,132.33, 129.76, 129.33, 126.21, 123.20, 122.08, 117.12, 116.79, 116.37,109.73, 95.35, 67.25, 58.60, 56.16, 55.59, 26.70, 24.93.

Compound 10.

A round bottom flask was charged with compound 9 (50 mg, 0.088 mmol),Pd₂dba₃ (4.1 mg, 5 mol %), KOH (20.1 mg, 4 equiv.), ligand2-Di-tert-butylphosphino-2′,4′,6′-triisopropylbiphenyl (4.2 mg, 10% mol)and filled with argon. A mixture of 1,4-dioxane (2 mL) and H₂O (1 mL)was added. The reaction was stirred at 80° C. for 6 hrs. Most of thesolvent was removed and directly purified by column chromatography(DCM/MeOH 50:3 with 0.1% acetic acid), product was obtained as acid salt(45 mg, 90%). ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.60 (d, 2H, J=8.8 Hz),7.56 (d, 1H, J=2.0 Hz), 7.26 (d, 1H, J=8.8 Hz), 7.01 (dd, 1H, J=2.0 Hz,8.8 Hz), 6.83-6.89 (m, 6H), 5.26 (s, 2H), 4.18 (t, 2H, J=5.6 Hz), 3.45(s, 3H), 3.02 (t, 2H, J=5.6 Hz), 2.84 (broad peak, 4H), 1.94 (s, 3H),1.65-1.70 (m, 4H), 1.48-1.50 (m, 2H); ¹³C-NMR (Acetone-d₆, 100 MHz):172.86, 158.54, 156.36, 154.66, 152.66, 140.08, 137.09, 129.79, 129.55,128.31, 124.24, 122.52, 117.02, 116.64, 116.47, 116.43, 109.84, 95.42,65.64, 57.54, 56.10, 54.79, 25.18, 23.77, 20.96.

Compound 12.

Compound 10 (40 mg, 0.07 mmol) was dissolved in DCM (20 mL) and washedwith saturated NaHCO₃ (2×10 mL), organic phase was separated,concentrated and dried under high vacuum for 1 hr. The residue wasdissolved in DCM (2 mL), DIPEA (116 μL, 0.7 mmol), DMAP (8.5 mg, 0.07mmol), compound 11 (16.9 mg, 0.1 mmol) and EDCI (27 mg, 0.14 mmol) wereadded successively. The reaction mixture was stirred at r.t. for 3 hrs,diluted with DCM (20 mL), washed with aqueous NaHCO₃ (10 mL) and H₂O (10mL), organic phase separated and concentrated. The residue was purifiedby column chromatography (ethyl acetate/MeOH 30:1), product was obtainedas slightly yellow oil (30 mg, 65%). ¹H-NMR (Acetone-d₆, 400 MHz): δ7.81-7.83 (m, 2H), 7.60 (d, 1H, J=2.0 Hz), 7.31 (d, 1H, J=8.8 Hz),7.18-7.21 (m, 2H), 7.03 (dd, 1H, J=8.8 Hz, 2.0 Hz), 6.84-6.92 (m, 4H),5.27 (s, 2H), 4.86 (s, 2H), 4.00 (t, 2H, J=6.0 Hz), 3.46 (s, 3H), 2.65(t, 2H, J=6.0 Hz), 2.45 (broad peak, 4H), 1.58-1.61 (m, 2H), 1.49-1.54(m, 4H), 1.35-1.40 (m, 4H); ¹³C-NMR (Acetone-d₆, 100 MHz): 171.46,156.82, 155.43, 152.23, 151.15, 141.89, 137.74, 130.92, 129.45, 129.12,126.70, 123.10, 123.01, 117.10, 116.75, 116.38, 109.78, 95.39, 76.53,67.31, 58.66, 56.15, 55.65, 26.76, 24.98, 22.35, 15.36.

Compound 13.

Compound 12 (30 mg, 0.046 mmol) was dissolved in a mixture of DCM (2 mL)and EtOH (0.5 mL), TsOH.H₂O (20 mg, 0.11 mmol) was added. The reactionmixture was stirred at 40° C. until all the starting material wasconsumed. After dilution with DCM (25 mL) and washed with saturatedNaHCO₃ (2×5 mL), organic phase was separated and concentrated. Theresidue was purified by column chromatography (DCM/MeOH 25:2), productwas obtained as slightly yellow foam (16 mg, 58%). ¹H-NMR (THF-ds, 400MHz): δ 7.43 (d, 2H, J=8.8 Hz), 7.15-7.17 (m, 2H), 7.08 (d, 2H, J=8.8Hz), 6.85 (d, 2H, J=9.2 Hz), 6.78 (d, 2H, J=9.2 Hz), 6.73 (dd, 1H, J=2.0Hz, 8.8 Hz), 4.76 (s, 2H), 4.00 (t, 2H, J=6.0 Hz), 2.70 (t, 2H, J=6.0Hz), 2.51 (broad peak, 4H), 1.51-1.59 (m, 6H), 1.38-1.43 (m, 4H);¹³C-NMR (THF-d₈, 100 MHz): 171.04, 157.50, 155.36, 152.52, 150.98,142.16, 138.22, 131.34, 128.83, 127.76, 125.04, 123.19, 122.62, 117.02,116.21, 115.57, 108.61, 76.32, 68.22, 58.68, 55.76, 26.72, 24.92, 22.44,15.19.

Compound 14.

4′FDMA (155 mg, 0.33 mmol), DIPEA (330 μL, 2.0 mmol), DMAP (30 mg, 0.25mmol), compound 14 (82 mg, 0.51 mmol) were dissolved in DCM (5 mL), EDCI(134 mg, 0.7 mmol) was added to this solution. The reaction was stirredat r.t. overnight. Reaction mixture was diluted with ethyl acetate (50mL), washed with water and concentrated, the residue was purified bycolumn chromatography (DCM/MeOH 25:1), product was obtained as whitefoam (151 mg, 74%). ¹H-NMR (Acetone-d₆, 400 MHz): δ 7.82-7.85 (m, 2H),7.78 (d, 1H, J=1.6 Hz), 7.41 (d, 1H, J=8.8 Hz), 7.18-7.23 (m, 2H), 7.13(dd, 1H, J=8.4 Hz, 2.0 Hz), 6.85-6.92 (m, 4H), 4.89 (s, 2H), 4.03 (t,2H, J=6.0 Hz), 2.69 (t, 2H, J=6.0 Hz), 2.49 (broad peak, 4H), 1.62-1.65(m, 2H), 1.50-1.55 (m, 4H), 1.38-1.41 (m, 4H); ¹³C-NMR (Acetone-d₆, 100MHz): δ 171.73, 164.54, 162.09, 155.46, 152.17, 149.59, 141.51, 136.81,132.65, (130.51, 130.43), (129.19, 129.16), 122.88, 120.38, 117.11,116.83, 116.61, 116.46, 76.54, 67.16, 58.53, 55.56, 26.59, 24.84, 22.40,15.43.

Example 13 Pharmacokinetics of SEMs in Mice

SEM prodrugs have been prepared using standard, scalable organicchemistry techniques. SEMs with antithrombotic properties will beprepared using the strategy described above for NO-SERMs (e.g.,compounds 8, 13, and 14). Metabolic stability will be verified in humanliver and human intestinal microsomal incubations, and plasma, with orwithout esterase (PLE). SEMs with high stability, minimal oxidativemetabolites, and efficient enzymic bioactivation to TTC-352 [i.e.,3-(4-fluorophenyl)-2-(4-hydroxyphenoxy)benzo[b]thiophen-6-ol]will beadvanced to pharmacokinetic studies.

Ovariectomized (OVX) nude 4- to 6-week-old athymic mice will beadministered TTC-352 and no more than 3 SEM prodrugs and/or NO-SEMs bygavage in ethanol using a vehicle of propylene glycol/carboxymethylcellulose at a single dose of 4.4 μmol (equivalent to 1.5 mg/day used inxenograft studies). Blood samples will be collected at 20 min, 2 h, and6 h after treatment, using EDTA as an anticoagulant. Plasma will beseparated from whole blood by centrifugation at 4° C. and samples willbe immediately analyzed for prodrug and drug using LC-MS/MS.

Example 14 Efficacy of SEM Prodrugs and NO-SEMs on TAM-ResistantXenograft Tumor Growth

T47D-Tam1 and T47D/PKCα xenograft tumor models will be used to assessthe efficacy of 2 SEM treatments at 2 doses (4.4 μmol and 0.44 μmol/day,based upon data shown in FIG. 11). Tumors will be grown in OVX athymicmice and treated once tumor volume reaches approximately 0.5 cm². Theresponse to SEMs and/or NO-SEMs will be compared to the standard of care(e.g., tamoxifen).

Example 15 Mechanism of Action Confirmation

For tumors treated by SEMs and/or NO-SEMs, PKCα, and ER biomarkers willbe assessed by immunohistochemistry. Anticoagulant activity will bemeasured ex vivo.

It is understood that the foregoing detailed description andaccompanying examples are merely illustrative and are not to be taken aslimitations upon the scope of the invention, which is defined solely bythe appended claims and their equivalents.

Various changes and modifications to the disclosed embodiments will beapparent to those skilled in the art. Such changes and modifications,including without limitation those relating to the chemical structures,substituents, derivatives, intermediates, syntheses, compositions,formulations, or methods of use of the invention, may be made withoutdeparting from the spirit and scope thereof.

Example 16 Synthesis of a Compound of Formula (I)

A compound of formula (I), wherein A₁ is halogen, A₂ is —OR₂, R₂ ishydrogen, X₁ is —O—, A₃ is aryl, and A₄ is —OR₃, wherein R₃ is describedabove, may be prepared according to the methods illustrated in Scheme 1,2 or 3.

A compound of formula (Ic) or formula (II) may be prepared according tothe methods illustrated in Scheme 4, 5, or 6.

The compound3-(4-fluorophenyl)-2-(4-hydroxyphenoxy)benzo[b]thiophen-6-ol (TTC-352)can be prepared as described above in Scheme 7 and the followingexamples.

1-(4-Fluorophenyl)-2-(3-methoxyphenylsulfanyl)ethanone

3-Methoxybenzenethiol (1 g, 7.1 mmol) was added in one portion to afreshly prepared solution of 7.5 mL of ethanol, 3 mL of water, and 470mg of KOH (8.4 mmol). The solution was cooled to 5-10° C. A solution of2-bromo-1-(4-fluorophenyl)ethanone (1.54 g, 7.1 mmol) in 2.5 mL of EtOAcwas added to this solution at a rate such that the temperature did notexceed 25° C., and the reaction mixture was allowed to stir overnight atroom temperature. The solvents were removed under reduced pressure, andthe residue was partitioned between water and ethyl acetate. The aqueouslayer was isolated and extracted several times with ethyl acetate, andthe combined extracts were washed with consecutive portions of 10% HCl,water, saturated NaHCO3, and water before being dried over anhydrousNa2SO4. After concentration in vacuo to an oil, the crude product waspurified by flash chromatography [SiO2, hexane/ethyl acetate (10:1,v/v)] to give 1.4 g (74%) desired product. 1H NMR (300 MHz, DMSO-d6) δ3.73 (s, 3H), 4.67 (s, 3H), 6.75 (m, 1H), 6.91 (m, 2H), 7.18 (t, J) 8.2Hz, 1H), 7.34 (t, J) 8.9 Hz, 2H), 8.12 (q, J) 8.9 Hz, 2H).

3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene

A flask was charged with1-(4-Fluorophenyl)-2-(3-methoxyphenylsulfanyl)ethanone and BF₃.OEt₂under Argon atmosphere at room temperature. The reaction mixture wasstirred until starting material was consumed as monitored by TLC. Thereaction mixture was poured into saturated NaHCO₃/ice water, stirred 30min, and extracted with dichloromethane. The crude product was purifiedby flash chromatography [SiO2, hexane/dichloromethane (20:1, v/v)]. Thecombined fractions from column was recrystalized to get 50% pureproduct. 1H NMR (400 MHz, CDCl₃) δ (ppm) 3.89 (s, 3H), 7.02 (dd, 8.9 Hz,2.2 Hz 1H), 7.15 (m, 3H), 7.37 (d, 2.2 Hz 1H), 7.52 (m, 2H), 7.71 (d,8.9 Hz, 1H), ¹³C NMR (100 MHz, CDCl₃) δ (ppm) 55.62, 105.24, 114.52,115.50, 115.71, 120.67, 123.30, 130.09, 130.17, 131.96, 132.17, 136.62,142.07, 157.56, 162.31.

2-bromo-3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene

N-Bromoacetamide (1.46 g, 10.5 mmol) in 10 mL of ethanol was addeddropwise to a solution of 3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene(2.58 g, 10 mmol) in 300 mL of CH₂Cl₂ and 20 mL of ethanol at roomtemperature. After the mixture was stirred for 1 h, the solvent wasremoved in vacuo. Next, the residue was titrated with ethanol andfiltered to give 3.0 g (89%) of desired product. ¹H NMR (400 MHz, CDCl₃)δ (ppm) 3.898 (s, 3H), 6.94 (dd, 8.9 Hz, 2.3 Hz 1H), 7.24 (m, 3H), 7.38(d, 8.9 Hz 1H), 7.45 (m, 2H)), ¹³C NMR (100 MHz, CDCl₃) δ (ppm) 55.27,104.1, 109.47, 114.20, 115.14, 115.36, 122.99, 129.57, 131.23, 131.31,132.29, 135.28, 140.61, 157.36, 162.05.

2-bromo-3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene 1-oxide

Trifluoacetic acid (13 mL) was added dropwise to a solution of2-bromo-3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene (2.4 g, 7 mmol) in13 mL of anhydrous CH₂Cl₂. After the mixture was stirred for 5 min, H₂O₂(1.0 mL, 7 mmol, 30% aqueous solution) was added dropwise, and theresulting mixture was stirred for 2 h at room temperature. Sodiumbisulfite (0.3 g) was added to the solution followed by 5 mL of water.The mixture was stirred vigorously for 30 min and then concentrated invacuo. The residue was partitioned between CH₂Cl₂ and saturated aqueousNaHCO₃ solution (50 mL each). The layers were separated, and the organiclayer was washed with consecutive portions of water, saturated NaHCO₃,and water, and then dried over anhydrous Na₂SO₄ and concentrated invacuo; the residue was titrated with diethylether and filtered to give2.1 g (84%) of desired product.

3-(4-fluorophenyl)-6-methoxy-2-(4-methoxyphenoxy)benzo[b]thiophene1-oxide

NaH (237 mg, 9.9 mmol, 60% dispersion in mineral oil) was added to asolution of 4-methoxyphenol (1.31 g, 59 mmol) in 25 mL of anhydrous DMFat room temperature. After the mixture was stirred for 15 min,2-bromo-3-(4-fluorophenyl)-6-methoxybenzo[b]thiophene 1-oxide (2 g, 5.7mmol) was added in small portions. After the mixture was stirred for 1h, ethyl acetate and water were added, and the organic layer was washedseveral times with water and then dried over Na₂SO₄. The residue wastitrated with hexane/ethyl acetate (10:1, v/v) and filtered to yield2.47 g (89%) of desired product.

3-(4-fluorophenyl)-6-methoxy-2-(4-methoxyphenoxy)benzo[b]thiophene

LiAlH₄ (0.27 g, 7.2 mmol) was added in small portions to a solution of3-(4-fluorophenyl)-6-methoxy-2-(4-methoxyphenoxy)benzo[b]thiophene1-oxide (2.37 g, 4.8 mmol) in 180 mL of anhydrous THF under argon at 0°C. After the mixture was stirred for 30 min, the reaction was quenchedby the slow addition of 4 mL of 2.0 MNaOH. The mixture was stirredvigorously for 30 min, and a minimal amount of 2.0 M NaOH was added todissolve salts. The mixture was then partitioned between water andethanol/ethyl acetate (1:9, v/v). The aqueous layer was isolated andthen extracted several times with ethanol/ethyl acetate (1:9, v/v). Theorganic layers were combined, dried over anhydrous Na2SO4, concentratedin vacuo to an oil, and then purified by flash chromatography to give1.2 g (67%) of desired product.

3-(4-fluorophenyl)-2-(4-hydroxyphenoxy)benzo[b]thiophen-6-ol (TTC-352)

3-(4-fluorophenyl)-6-methoxy-2-(4-methoxyphenoxy)benzo[b]thiophene (1.5g, 2.9 mmol) was dissolved in 150 mL of anhydrous CH2Cl2 and cooled to0° C. BBr₃ (1.0 M in CH2Cl2, 11.6 mL, 11.6 mmol) was added to thissolution followed by stirring at 0° C. for 4 h. The reaction wasquenched by saturated NaHCO3 (100 mL) and cooled to 0° C. The aqueouslayer was isolated and extracted with methanol/ethyl acetate (5:95, v/v)(3_100 mL). The organic extracts were combined, dried over anhydrousNa2SO4, concentrated in vacuo, and then purified by flash chromatography[SiO2, CH2Cl2/MeOH (8:1, v/v)] to obtain 1.1 g (75%) desired product. ¹HNMR (400 MHz, acetone d6) δ (ppm) 6.84 (m, 2H), 6.97 (dd, 8.9 Hz, 2.2Hz, 1H), 7.02 (m, 2H), 7.24 (m, 2H)), 7.28 (d, 2.2 Hz, 1H), 7.47 (d, 8.9Hz, 1H), 7.61 (m, 2H)¹³C NMR (100 MHz acetone d6) δ (ppm) 162.84,155.94, 155.02, 153.81, 152.35, 135.21, 132.28, 132.20, 130.90, 130.32,123.70, 120.81, 119.67, 119.66, 116.95, 116.95, 116.39, 116.17, 115.52,109.00.

4-((3-(4-fluorophenyl)-6-methoxybenzo[b]thiophen-2-yl)oxy)phenol(Monomethoxyl-TTC-352)

Made from general procedure as TTC-352. ¹H NMR (400 MHz, Acetone d6)δ=7.62 (m, 2H), 7.52 (d, 8.9 Hz, 1H), 7.41 (d, 2.3 Hz, 1H), 7.25 (t, 8.9Hz, 2H), 7.02 (dd, 9.0 Hz, 2.6, 3H), 6.83 (d, 9.0 Hz, 2H), 3.86 (s, 3H).¹³C NMR (100 MHz acetone d6) δ (ppm) 161.95, 157.52, 154.09, 153.58,151.34, 134.15, 131.34, 131.26, 130.71, 129.25, 122.60, 119.70, 118.79,118.79, 116.00, 115.91, 115.46, 115.24, 114.20, 105.89, 55.04.

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What is claimed is:
 1. A compound having a formula of (II-i):

or a pharmaceutically acceptable salt thereof, wherein A₁ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₁; A₂ isselected from the group consisting of halogen and trifluoromethyl; A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl; A₄ is selected from thegroup consisting of hydrogen, halogen, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted, and—OR₃; R₁ is selected from the group consisting of —SO₃R^(x1) and—PO₃R^(y1)R^(z1); R₃ is selected from the group consisting of hydrogen,alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), and —C(═O)R^(a); R^(x1), R^(y1) andR^(z1), at each occurrence, are independently selected from the groupconsisting of hydrogen and a pharmaceutically acceptable cation; andR^(a) is alkyl or —OH.
 2. A compound selected from the group consistingof:(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(4-(trifluoromethyl)phenyl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(4-(trifluoromethyl)phenyl)methanone;cyclopropyl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;cyclopropyl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)(pyridin-4-yl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(pyridin-4-yl)methanone;1-(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)-2-methylpropan-1-one;1-(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)-2-methylpropan-1-one;(4-ethynylphenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-ethynylphenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)(p-tolyl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;(4-(1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)phenyl)(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;(3r,5r,7r)-adamantan-1-yl(2-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-3-yl)methanone;and(3r,5r,7r)-adamantan-1-yl(3-(4-fluorophenyl)-6-hydroxybenzo[b]thiophen-2-yl)methanone;or a pharmaceutically acceptable salt thereof.
 3. A compound having aformula of (II-ii):

or a pharmaceutically acceptable salt thereof, wherein A₁ is selectedfrom the group consisting of halogen, trifluoromethyl, and —OR₁; A₂ isselected from the group consisting of halogen and trifluoromethyl; A₃ isselected from the group consisting of alkyl, aryl, cycloalkyl,cycloalkylalkyl, heterocycle, and heteroaryl; A₄ is selected from thegroup consisting of hydrogen, halogen, alkyl, cyano, trifluoromethyl,aryl-heteroaryl wherein the aryl is substituted or unsubstituted, and—OR₃; R₁, R₂ and R₃ are each independently selected from the groupconsisting of hydrogen, alkyl, —SO₃R^(x1), —PO₃R^(y1)R^(z1), and—C(═O)R^(a); R^(x1), R^(y1) and R^(z1), at each occurrence, areindependently selected from the group consisting of hydrogen and apharmaceutically acceptable cation; and R^(a) is alkyl or —OH.